Estimation Of Cell Volume Research Articles

Quantification of bacterial shape using moment invariants enables distinguishing populations during cellular plasmolysis

The analysis of geometrical cell shape is fundamental to understand motility, development, and responses to external stimuli. The moment invariants framework quantifies cellular shape and size, although its applicability has not been explored for rod-shaped bacteria. In this work, we use moment invariants to evaluate the extent of cell shape change (projected area and volume) during plasmolysis, as Escherichia coli cells are subjected to hyperosmotic shock. The characteristic cell size descriptors width, length and area show systematic decrease as external salt (NaCl) conditions increase—except for high salt, where a small population of cells shows evidence of membrane rupture. We use these two-dimensional results to estimate cell volume during plasmolysis, finding a minimum volume that is not reduced further with increase in salt concentration. Next, we computed elongation and dispersion, metrics that quantify how cell shape is stretched out or differs from an ellipse, respectively. For dispersion, we observe the development of a long tail for the distribution at high salt. Moreover, the use of elongation-dispersion plots enables distinction of plasmolyzed and normal cells despite the presence of broad distributions. Altogether, a protocol is provided to evaluate bacterial shape, highlighting a set of metrics that help distinguish among bacterial populations.•Moment invariants enable quantitative description of bacterial morphology in two dimensions, and estimation of volume•We apply the moment invariants framework to describe changes in bacterial shape during plasmolysis•The proposed methodology shows suitability to distinguish among cellular populations.

Gene-dosage- and sex-dependent differences in the prodromal-Like phase of the F344tgHD rat model for Huntington disease.

In Huntington disease (HD) the prodromal phase has been increasingly investigated and is currently in focus for early interventional treatments. Also, the influence of sex on disease progression and severity in patients is under discussion, as a sex-specific impact has been reported in transgenic rodent models for HD. To this end, we have been studying these aspects in Sprague Dawley rats transgenic for HD. Here, we took up on the congenic F344tgHD rat model, expressing a fragmented Htt construct with 51 CAG repeats on an inbred F344 rat background and characterized potential sexual dimorphism and gene-dosage effects in rats during the pre-symptomatic phase (1-8 months of age). Our study comprises a longitudinal phenotyping of motor function, emotion and sensorimotor gating, as well as screening of metabolic parameters with classical and automated assays in combination with investigation of molecular HD hallmarks (striatal cell number and volume estimation, appearance of HTT aggregates). Differences between sexes became apparent during middle age, particularly in the motor and sensorimotor domains. Female individuals were generally more active, demonstrated different gait characteristics than males and less anxiolytic-like behavior. Alterations in both the time course and affected behavioral domains varied between male and female F344tgHD rats. First subtle behavioral anomalies were detected in transgenic F344tgHD rats prior to striatal MSN cell loss, revealing a prodromal-like phase in this model. Our findings demonstrate that the congenic F344tgHD rat model shows high face-validity, closely resembling the human disease's temporal progression, while having a relatively low number of CAG repeats, a slowly progressing pathology with a prodromal-like phase and a comparatively subtle phenotype. By differentiating the sexes regarding HD-related changes and characterizing the prodromal-like phase in this model, these findings provide a foundation for future treatment studies.

Evaluacija performansi Veri-Q hemoglobinskog merača za procenu hemoglobina i hematokrita na mestu pružanja pomoći

Abstract: Introduction: Haemoglobin (Hb) and packed cell volume (PCV) are tests used in the assessment of anaemia. The Veri-Q haemoglobin meter is a new device in the point-of-care market used for the quantitative measurement of haemoglobin and packed cell volume. This study aimed to evaluate the performance of the Veri-Q haemoglobin meter for the assessment of haemoglobin and packed cell volume. Materials and Methods: Haemoglobin and packed cell volume estimations of one hundred and eleven participants were determined using the Veri-Q Red haemoglobin meter and statistically compared with those obtained from the conventional method (gold standard). Results: One hundred and eleven undergraduate students participated in this study, of whom 72 (64.9%) were females and 39 (35.1%) were males. The mean haemoglobin values calculated using the Veri-Q haemoglobin meter (11.97 ± 1.95 g/dl) were significantly lower than the values obtained with the conventional method (12.35 ± 1.66 g/dl) (t = 4.7549; p = 0.0001). Similar results were obtained with packed cell volume estimation. The performance indices of the Veri-Q haemoglobin meter were as follows: sensitivity (75.41% for Hb and 77.05% for PCV), specificity (28.0% for Hb and PCV), positive predictive value (PPV) (56.1% for Hb and 56.63% for PCV), negative predictive value (48.28% for Hb and 50% for PCV). The prevalence of anaemia in this study was 54.05%. Positive Likelihood Ratio (1.05 for Hb and 1.07 for PCV), Negative Likelihood Ratio (0.88 for Hb and 0.82 for PCV), Accuracy (54.05% for Hb and 54.95% for PCV). The Receiver Operative Characteristics (ROC) Area Under Curve (AUC) was 0.996 for Hb and 0.984 for PCV. A significant but weak positive correlation was found to exist between haemoglobin estimations using conventional and Veri-Q methods (r = 0.3013, P = 0.01); PCV (R = 0.2512, P = 0.07). The sensitivity of the device can be described as moderate while the specificity is low. The accuracy of the device is just average. Conclusion: The Veri-Q haemoglobin meter demonstrates an average level of accuracy and a high AUC, making it potentially useful for field epidemiological studies.

Comparison of two Phaeodactylum tricornutum ecotypes under nitrogen starvation and resupply reveals distinct lipid accumulation strategies but a common degradation process.

Phaeodactylum tricornutum is a model species frequently used to study lipid metabolism in diatoms. When exposed to a nutrient limitation or starvation, diatoms are known to accumulate neutral lipids in cytoplasmic lipid droplets (LDs). Those lipids are produced partly de novo and partly from the recycle of plastid membrane lipids. Under a nitrogen resupply, the accumulated lipids are catabolized, a phenomenon about which only a few data are available. Various strains of P. tricornutum have been isolated around the world that may differ in lipid accumulation patterns. To get further information on this topic, two genetically distant ecotypes of P. tricornutum (Pt1 and Pt4) have been cultivated under nitrogen deprivation during 11 days followed by a resupply period of 3 days. The importance of cytoplasmic LDs relative to the plastid was assessed by a combination of confocal laser scanning microscopy and cell volume estimation using bright field microscopy pictures. We observed that in addition to a basal population of small LDs (0.005 μm3 to 0.7 μm3) present in both strains all along the experiment, Pt4 cells immediately produced two large LDs (up to 12 μm3 after 11 days) while Pt1 cells progressively produced a higher number of smaller LDs (up to 7 μm3 after 11 days). In this work we showed that, in addition to intracellular available space, lipid accumulation may be limited by the pre-starvation size of the plastid as a source of membrane lipids to be recycled. After resupplying nitrogen and for both ecotypes, a fragmentation of the largest LDs was observed as well as a possible migration of LDs to the vacuoles that would suggest an autophagic degradation. Altogether, our results deepen the understanding of LDs dynamics and open research avenues for a better knowledge of lipid degradation indiatoms.

A new hyperelastic lookup table for RT-DC.

Real-time deformability cytometry (RT-DC) is an established method that quantifies features like size, shape, and stiffness for whole cell populations on a single-cell level in real-time. A lookup table (LUT) disentangles the experimentally derived steady-state cell deformation and the projected area to extract the cell stiffness in the form of the Young's modulus. So far, two lookup tables exist but are limited to simple linear material models and cylindrical channel geometries. Here, we present two new lookup tables for RT-DC based on a neo-Hookean hyperelastic material numerically derived by simulations based on the finite element method in square and cylindrical channel geometries. At the same time, we quantify the influence of the shear-thinning behavior of the surrounding medium on the stationary deformation of cells in RT-DC and discuss the applicability and impact of the proposed LUTs regarding past and future RT-DC data analysis. Additionally, we provide insights about the cell strain and stresses, as well as the influence resulting from the rotational symmetric assumption on the cell deformation and volume estimation. The new lookup tables and the numerical cell shapes are made freely available.

Stimulation of calcium influx and CK1α by NF-κB antagonist [6]-Gingerol reprograms red blood cell longevity.

Chemotherapy-induced anemia (CIA) is a major obstacle in cancer management. Although the mechanisms governing CIA are poorly understood, recent efforts have identified suicidal erythrocyte (red blood cell, RBC) death as a possible cause of CIA. [6]-Gingerol (GNG), a polyphenol extracted from Zingiber officinale plant, exhibits a wide array of biological activities including antimicrobial, antioxidant, anti-inflammatory, immunomodulatory, and anticancer activities, in vitro and in vivo. However, the potential toxicity of GNG to human RBCs remains unexplored. RBCs from heparinized blood were isolated by centrifugation and exposed to antitumor concentrations (10-100µM) of GNG for 24hr at 37°C. Hemolysis was calculated from hemoglobin leakage in the supernatant (λmax =405nm), while cytofluorometric analysis of eryptosis employed Annexin-V-FITC to detect phosphatidylserine (PS) exposure, forward scatter (FSC) to estimate cell volume, Fluo4/AM to measure calcium activity, and H2 DCFDA to assess oxidative stress. Moreover, zVAD(OMe)-FMK, SB203580, necrostatin-2, staurosporin, and D4476 were used to identify signaling pathways responsive to GNG. GNG induced significant hemolysis at 100µM, independently of extracellular calcium, and increased Annexin-V-FITC fluorescence that was thoroughly abrogated without extracellular calcium. GNG also enhanced Fluo4 fluorescence and reduced FSC, but had no significant effect on DCF fluorescence. Importantly, the presence of D4476 significantly attenuated GNG-induced hemolysis. In conclusion, GNG stimulates premature RBC death characterized by loss of membrane asymmetry, elevated cytosolic calcium, cell shrinkage, and casein kinase 1α activation. Blocking the activity of calcium channels or CK1α may, therefore, ameliorate the toxic effects of GNG on RBCs. PRACTICAL APPLICATIONS: This report presents a safety assessment of GNG as a chemotherapeutic agent and highlights the novel toxicity of GNG to human RBCs. Our findings provide novel insights that may lead to more efficient utilization of GNG in chemotherapy. Specifically, our data revealed the involvement of calcium channels and casein kinase 1α in mediating GNG-induced premature RBC death, and, therefore, inverse agonists or inhibitors of either pathway may be used as pharmaceutical adjuvants to attenuate the toxic effects of GNG.

Effect of discriminate and indiscriminate use of oxytetracycline on residual status in broiler soft tissues.

Aim:This study aimed to evaluate the effects of discriminate and indiscriminate use of oxytetracycline on hematological parameters, residual status in soft tissue of broiler and of thermal effect on oxytetracycline residual status.Materials and Methods:Eighteen, day-old male broiler chickens were purchased and were divided into three different groups (control group, discriminate group, and indiscriminate group). The control group received no antibiotics. The discriminate group received oxytetracycline 1 g/L drinking water for 5 consecutive days, and 10 days’ withdrawal period was maintained before sacrifice. The indiscriminate group received oxytetracycline 1 g/L drinking water till the sacrificed day. Blood samples were collected before sacrificing for hematological analysis. After sacrificing liver, kidney, spleen, and muscle samples were collected for analysis of oxytetracycline residues in raw soft tissues. Since meat is used to cook by traditional method in Bangladesh before consumption that is why positive meat samples were cooked by traditional cooking method to evaluate the thermal effect on oxytetracycline residual status as well. Thin-layer chromatography (TLC) was done for screening of oxytetracycline residues in soft tissues.Results:Mean differences of total erythrocyte count (million/mm3), hemoglobin estimation (gm%), and packed cell volume (%) estimation were not statistically significant among the groups. TLC analysis of raw samples showed 100% positive results of all samples collected from the indiscriminate group. In contrast, samples collected from the discriminate group were negative for oxytetracycline residues. In the control group, all samples were negative for oxytetracycline residue. There was a significant (p<0.05) relationship of oxytetracycline residues among three different groups for liver, kidney, spleen, and muscle samples. Positive liver and muscle samples from the indiscriminate group were subjected to thermal treatment by traditional cooking method of Bangladesh. Oxytetracycline residues had found in cooked meat, liver, and juice part, suggesting that antibiotic residues disseminated to juice part from flesh part after cooking.Conclusion:Evidence suggests that proper maintenance of withdrawal period would minimize oxytetracycline residues in broiler soft tissues, whereas antibiotics retained in soft tissues of broiler in case of indiscriminate use. Traditional cooking does not change oxytetracycline residual status in edible tissues. Therefore, awareness regarding the proper maintenance of withdrawal period after antibiotic treatment of broiler is one of the best strategies which may positively reduce the risk of antimicrobial drugs residue in meat.

Three-dimensional analysis of inter-and intraspecific variation in ontogenetic growth trajectories of planktonic foraminifera

Abstract Planktonic foraminifera grow by adding chambers onto their tests, modifying their surface area, volume, porosity, and ornamentation with each addition. This process results in a wide variety of adult morphologies and preserves a record of the early morphology. With high-resolution three-dimensional imaging techniques, precise measurements can be made on previously inaccessible parameters like surface area and volume of the test interior. Here, we used three-dimensional images (microCT scans) of extant planktonic foraminifera from plankton tows and sediment traps to measure and compare the variation in chamber and whole test volume within and between seven morphospecies. By incorporating data from Caromel et al., (2015), we were able to explore the extent of interspecific variation in growth (i.e., total volume) with each chamber addition from a total of 12 extant species. All species measured approximately adhered to log-linear growth trajectories through the majority of their ontogeny. Species exhibited characteristic deviations from the log-linear growth trajectory and were grouped into three categories on the basis thereon. The volume of the first chamber—the proloculus—varied by more than two orders of magnitude. However, subtle differences in relative growth rates could make up for the initial variation proloculus size, such that the correlation between proloculus size and biovolume decreased through ontogeny. Intraspecific variation in proloculus size, cavity volume at the final five chamber stages, and terminal calcite volume was pronounced in the seven morphospecies investigated and attributed environmental and ontogenetic differences among individuals. Finally, we generated linear models for estimating cell volume and calcium carbonate test volume from linear measurements (major and minor axis) that are more accurate than the spherical equivalents used previously.

A Prospective Study: Is Routine Pre-operative Urinalysis Mandatory in Ambulatory Operations in Children

There are controversies regarding pre-operative routine investigations in ambulatory operations in the paediatric age group. Routinely, patients’ packed cell volume (PCV), and genotype were the investigations done for children before surgery, but somewhere along the line, the anaesthesiologists now demand for pre-operative routine urinalysis in children going for ambulatory operations to help detect occult urinary tract diseases. The aim and objectives was to find out the benefits of doing routine preoperative urinalysis and whether it was really necessary in children undergoing ambulatory operations in Lagos, Nigeria”? It was a one year prospective study, carried out from the 1st of September, 2018 to the 31st of August, 2019 in the Lagos State University Teaching Hospital, Ikeja, Lagos, Nigeria. Eighty four children, between the ages of 0 - 12 years, slated for ambulatory operations were mandated to do routine pre-operative urinalysis, routine packed cell volume estimation, and genotype studies. All the children were operated under general anaesthesia. Eighty four children were recruited, comprising of 72 (85.7%) males and 12 (14.3%) females with a male: female ratio of 6:1. The patients’ age ranged from 0 to 12 years with a mean age of 5 ± 3 STD. An average, minimum and maximum weights of 18.9kg, 5.0kg, and 44.0kg respectively were recorded, while the average PCV was 34.4%, with a minimum of 26.9% and maximum of 48.0%. The results of urinalysis were normal in 82.1% and abnormal in about 17.9% participants. In the abnormal group, (proteinuria trace = 7, leucocytes = 4, nitrites = 2, proteins + nitrites = 1, and urobilirubin = 1). All study subjects underwent the procedures successfully with no record of cancellation of any procedure as a result of an abnormal urine finding. We conclude that routine pre-operative urinalysis should be done on all our paediatric ambulatory surgery patients with a view to detecting any abnormalities that may require a follow-up.

Practical implementation of the planar and spatial rotator in a complex tissue: the brain.

In neuroscience, application of widely used stereological local volume estimators, including the planar rotator, is challenged by the combination of a complex tissue organisation and an estimator requirement of either isotropic or vertical sections, i.e. randomly oriented tissue. The spatial rotator is applicable with any tissue orientation but is sensitive to projection artefacts. The challenge is thus to select the most appropriate method for individual analyses. In this study, agreement between estimates of mean cell volume acquired with the vertical planar and the spatial rotator is assessed for two brain regions with different types of cytoarchitecture (motor cortex and hippocampal cornu ammonis 1). The possibility of using the planar rotator in tissues cut in an arbitrary direction is explored and requirements for a theoretically unbiased result as well as histological considerations are provided. LAY DESCRIPTION: Cells may change volume both during disease and with advancing age. Assessment of the volume of individual cells can therefore serve as a useful indicator of general tissue state. Most available methods to estimate cell volume in tissue sections, however, require that the tissue analysed has random orientation. Particularly for complex tissues such as the brain this is a challenge as identification, delineation and subdivision of many brain areas rely heavily on the use of anatomical atlases where illustrations depict the tissue in a few well-known orientations. In this study, the practical application of two different methods for estimating mean cell volumes in tissues cut in a preferred orientation is evaluated. Requirements for the feasibility of cell volume estimation without random tissue orientation as well as histological considerations are provided.

Using video microscopy to improve quantitative estimates of protozoal motility and cell volume

The objective of this study was to apply digital imaging to improve quantification of rumen protozoal biomass and distinguish treatment differences in cell motility and volume among ruminal protozoa. Observations of protozoa in rumen fluid treated with essential oils (CinnaGar, CIN; Provimi North America, Brookville, OH) or an ionophore (monensin, MON) indicated possible cell shrinkage. We hypothesized that MON would decrease protozoal motility and interact with CIN on cell volume. In addition, we hypothesized that analysis of still frames from video of swimming protozoa would improve volume prediction accuracy. Flocculated rumen fluid was incubated in batch culture dosed with N-free feed only (control), MON, CIN, or a combination of MON+CIN. Samples were taken at 0, 3, or 6 h post-treatment and wet-mounted on a microscope fitted with a high-definition camera. At 3 h post-inoculation, there was a treatment interaction for average speed such that CIN attenuated the effect of MON, with treatment means of 243, 138, 211, and 183 µm/s for control, MON, CIN, and MON+CIN, respectively. At 6 h post-inoculation, MON decreased average speed by 79.2 µm/s compared with the main effect mean without MON. We measured both minimum and maximum diameters (depth and width, respectively) perpendicular to the longitudinal axis of swimming protozoa, yielding a 3-dimensional estimate of protozoal volume. The ellipsoid formula (4/3)πabc, where a = 1/2 length, b = 1/2 width, and c = 1/2 depth, was compared with previously published volume estimations using genera-specific coefficients (genera-specific coefficient × length × width2). Residuals (genera-specific coefficients - ellipsoid) were plotted against predicted (ellipsoid) and centered to the mean (Xi-x¯) to evaluate both mean and slope biases. For Entodinium spp., Y = 0.248 (±0.037) (Xi - 7.98 × 104) + 1.97 × 104 (±1.48 × 103); n = 100; r2 [coefficient of determination (squared correlation coefficient)] = 0.31, with significant slope and mean biases. For family Isotrichidae, Y = -0.124 (±0.068) (Xi - 2.54 × 106) - 1.21 × 104 (±4.86 × 104); n = 32; r2 = 0.10, where slope tended to be different from zero but with no mean bias. For Epidinium spp., Y = 0.375 (±0.056) (Xi - 2.45 × 105) + 6.65 × 104 (±0.28 × 104); n = 64; r2 = 0.43, with both mean and slope biases. The present regression analyses demonstrate that the genera-specific coefficient-based method more likely overestimates volume for Entodinium and Epidinium than for the teardrop-shaped Isotrichidae. Based on simulations derived from previous literature reporting treatments that depress protozoal populations or among-animal changes in protozoal population structures, our proposed ellipsoid method offers potential to advance the prediction of treatment effects on protozoal volume and to shift focus from the number of cells present to the diversity, function, and biomass of protozoa under various treatment conditions.

Estimation of diatom and dinoflagellate cell volumes from surface waters of the Northern Indian Ocean

Summary Phytoplankton samples collected from the Northern Indian Ocean (Bay of Bengal, northern Arabian Sea, and Dona Paula Bay Goa, west coast of India), were utilized to quantify changes in cell size, cell volume and carbon per cell of diatoms and dinoflagellates. The dataset from the Bay of Bengal also provides inter- and intra-annual variations (April 2008 to March 2010). The variations in cell size and volume were large in regions influenced by the riverine influx or terrigenous inputs. An interregional comparison of commonly available forms (8 species) points out that cell volumes are highest in the North Atlantic and lowest in the Mediterranean. The information provided will be useful in estimation of carbon biomass and biogeochemical studies.

Research of an algorithm for crystal lattice parameter identification based on the gradient steepest descent method

In the analysis of a crystalline substance, the problem of crystal lattice parameter identification is of a great interest. However, the existing methods for solving this problem, such as the Bravais cell parameters estimation method and Wigner-Seitz cell volume estimation method, do not provide the required level of accuracy. Aiming to address the problem of low identification accuracy, the paper proposes an algorithm for crystal lattice parameter identification based on the gradient steepest descent method. The study of the feasibility of the structure parameter identification is carried out using a large set of distorted lattices. The results obtained show a significant increase in the identification accuracy in comparison with the above-mentioned parameter identification methods.

Prevalence and predictors of placental malaria in human immunodeficiency virus-positive women in Nigeria.

Human immunodeficiency virus (HIV)-infected pregnant women have alterations in cellular and humoral immunity that increase the risks to placental malaria infection. This study aimed at determining the prevalence and predictors of placental malaria among HIV-positive women in Nigeria. It was a longitudinal cohort study of pregnant women receiving antenatal care at a tertiary hospital in Nigeria. Peripheral blood sample for packed cell volume estimation and placental blood sample for malaria parasite estimation were collected from each participant at a presentation in labor and upon delivery, respectively. The Prevalence of placenta malaria (68.6%) and anemia (66.7%) in HIV-positive women were significantly higher than the prevalence of placental malaria (35.3%) and anemia (44.1%) in HIV-negative control (P < 0.001 and P = 0.001 respectively). The employment status was the only sociodemographic factor significantly associated with the development of placental malaria in HIV-positive women (odds ratio: 21.60; 95% confidence interval: 7.1-66.2; P< 0.001). The prevalence of placental malaria is very high among HIV-positive women in Nigeria. Scaling up free distribution of insecticide treated nets in the short term and employment opportunities of HIV-positive women, in the long run, may reduce the prevalence of placental malaria in our population.

Triggering of Eryptosis, the Suicidal Erythrocyte Death by Mammalian Target of Rapamycin (mTOR) inhibitor Temsirolimus

Background/Aims: The mammalian target of rapamycin (mTOR) inhibitor temsirolimus is utilized for the treatment of malignancy. Temsirolimus is at least in part effective by triggering suicidal tumor cell death. The most common side effect of temsirolimus treatment is anemia. At least in theory, the anemia following temsirolimus treatment could result from stimulation of eryptosis, the suicidal erythrocyte death. Hallmarks of eryptosis include cell shrinkage and cell membrane scrambling with phosphatidylserine translocation to the erythrocyte surface. Signaling involved in the orchestration of eryptosis include increase of cytosolic Ca²⁺ activity ([Ca²⁺]_i), oxidative stress, ceramide, as well as activation of staurosporine and chelerythrine sensitive protein kinase C, SB203580 sensitive p38 kinase, D4476 sensitive casein kinase 1, and zVAD sensitive caspases. The purpose of the present study was to test whether temsirolimus influences eryptosis and, if so, to shed light on the signaling involved. Methods: Flow cytometry was employed to estimate cell volume from forward scatter, phosphatidylserine exposure at the cell surface from annexin-V-binding, [Ca²⁺]_i from Fluo3-fluorescence, reactive oxygen species (ROS) abundance from DCFDA dependent fluorescence, and ceramide abundance utilizing specific antibodies. Hemolysis was determined from hemoglobin concentration in the supernatant. Results: A 48 hours exposure of human erythrocytes to temsirolimus (5 – 20 µg/ml) significantly decreased forward scatter and significantly increased the percentage of annexin-V-binding cells. Temsirolimus significantly increased Fluo3-fluorescence, DCFDA fluorescence and ceramide abundance at the erythrocyte surface. The effect of temsirolimus on annexin-V-binding was significantly blunted but not abolished by removal of extracellular Ca²⁺ and by addition of staurosporine (1 µM) or chelerythrine (10 µM) but not significantly modified by addition of SB203580 (2 µM), D4476 (10 µM), or zVAD (10 µM). Chelerythrine (10 µM) further significantly blunted the effect of temsirolimus on DCFDA fluorescence but not ceramide formation. Removal of extracellular Ca²⁺ had no effect on temsirolimus induced ROS formation or ceramide abundance. Conclusions: Temsirolimus triggers eryptosis with cell shrinkage and phospholipid scrambling of the erythrocyte cell membrane, an effect at least in part due to Ca²⁺ entry, oxidative stress, ceramide and activation of staurosporine/Chelerythrine sensitive kinase(s).

Effects of Lugol’s iodine solution and formalin on cell volume of three bloom-forming dinoflagellates

Fixatives are traditionally used in marine ecosystem research. The bias introduced by fixatives on the dimensions of plankton cells may lead to an overestimation or underestimation of the carbon biomass. To determine the impact of traditional fixatives on dinoflagellates during short- and long-term fixation, we analyzed the degree of change in three bloom-forming dinoflagellates (Prorocentrum micans, Scrippsiella trochoidea and Noctiluca scintillans) brought about by Lugol’s iodine solution (hereafter Lugol’s) and formalin. The fixation effects were species-specific. P. micans cell volume showed no significant change following long-term preservation, and S. trochoidea swelled by approximately 8.06% in Lugol’s and by 20.97% in formalin as a percentage of the live cell volume, respectively. N. scintillans shrank significantly in both fixatives. The volume change due to formalin in N. scintillans was not concentration-dependent, whereas the volume shrinkage of N. scintillans cells fixed with Lugol’s at a concentration of 2% was nearly six-fold that in cells fixed with Lugol’s at a concentration of 0.6%–0.8%. To better estimate the volume of N. scintillans fixed in formalin at a concentration of 5%, we suggest that the conversion relationship was as follows: volume of live cell=volume of intact fixed cell/0.61. Apart from size change, damage induced by fixatives on N. scintillans was obvious. Lugol’s is not a suitable fixative for N. scintillans due to high frequency of broken cells. Accurate carbon biomass estimate of N. scintillans should be performed on live samples. These findings help to improve the estimate of phytoplankton cell volume and carbon biomass in marine ecosystem.

EFFECTS OF TULSI LEAF EXTRACT ON BODY WEIGHT GAIN IN BROILER PRODUCTION

The objective of this research was to evaluate the effect of tulsi leaf (Ocimum sanctum) extract supplementation in drinking water as a growth promoter in broiler chickens. A total of 40 Cobb-500 broiler chicks (day-old) were purchased from local hatchery (Nourish Poultry and Hatchery Ltd.). After seven days of acclimatization chicks were randomly divided into two groups, A (n=20) and B (n=20). The group A was kept as a control and not treated. The group B was supplemented with tulsi leaf extract with feed and water. Weekly observations were recorded for live body weight gain up to 5th weeks and hematological tests were performed at 35th day’s age of broiler to search for hematological changes between control (A) and treatment (B) groups. The initial body weight of groups A and B on 1st day of this experiment were 41.00±1.78 gm and 41.50±2.35 gm, respectively and after 35th day of experiment final body weight were 1640± 74.83 gm and 1920± 33.91 gm, respectively and economics of production were analyzed and found that net profit per broiler was Tk. 21.46 and Tk. 32.29, respectively. The treatment group B was recorded statistically significant (at 1% level) increased (16.97%) for live body weight than that of control group A. The hematological parameters total erythrocyte count (TEC), erythrocyte sedimentation rate (ESR), packed cell volume (PCV) and hemoglobin (Hb) estimation value of treatment group shows significant difference, while hemoglobin (Hb) estimation does not show significant difference from control group. The results suggest that better growth performance could be achieved in broilers supplemented with tulsi leaf extract.

Ritonavir-Induced Suicidal Death of Human Erythrocytes.

The antiviral drug ritonavir has been shown to trigger suicidal death or apoptosis of tumour cells and has thus been considered for the treatment of malignancy. In analogy to apoptosis of nucleated cells, erythrocytes may enter eryptosis, the suicidal erythrocyte death characterized by cell shrinkage and cell membrane scrambling with phosphatidylserine translocation to the erythrocyte surface. Triggers of eryptosis include Ca(2+) entry with increase in cytosolic Ca(2+) activity ([Ca(2+) ]i ), oxidative stress and ceramide. The present study explored whether and how ritonavir induces eryptosis. To this end, flow cytometry was employed to estimate cell volume from forward scatter, phosphatidylserine exposure at the cell surface from Annexin-V binding, [Ca(2+) ]i from Fluo3 fluorescence, abundance of reactive oxygen species (ROS) from 2',7'-dichlorodihydrofluorescein diacetate (DCFDA) fluorescence and ceramide abundance utilizing specific antibodies. As a result, a 48-hr exposure of human erythrocytes to ritonavir significantly increased the percentage of Annexin-V-binding cells (≥5 μg/ml), significantly decreased forward scatter (≥5 μg/ml), significantly increased Fluo3 fluorescence (20 μg/ml), slightly, but significantly increased DCFDA fluorescence (20 μg/ml) and slightly, but significantly increased ceramide abundance (20 μg/ml). The effect of ritonavir on Annexin-V binding was significantly blunted, but not fully abolished by the removal of extracellular Ca(2+) . In conclusion, ritonavir triggers erythrocyte shrinkage and phosphatidylserine translocation at the erythrocyte cell membrane, an effect in part due to the stimulation of Ca(2+) entry, oxidative stress and ceramide.

On the 3D reconstruction of diatom frustules: a novel method, applications, and limitations

Because of the importance of diatoms and the lack of information about their third dimension, a new method for the 3D reconstruction is explored, based on digital image correlation of several scanning electron microscope images. The accuracy of the method to reconstruct both centric and pennate (symmetrical and asymmetrical) diatoms was shown, independently of valve size and shape, and considering not only the general frustule morphology but also the intricate ornamentation. Several measurements were obtained, such as of the surface and projected areas and the valve volume. These results were validated by focused ion beam transverse cross section of one valve, and the quantitative results were compared with geometric models commonly applied. Furthermore, direct volume calculations based on 3D reconstructions have several advantages, such as it has higher taxonomic accuracy than the methods based on light microscopy; it solves the problems of the light halos; it allows the precise measurement of all linear dimensions, including the often neglected third dimension; natural samples can be measured directly; and it provides an exact estimate of cell volume, independently of its shape or alterations due to life cycle stage. This approach provides therefore a simple way to measure the morphological features of diatoms, even at a nanoscale, and can be applied to other microorganisms commonly illustrated by means of scanning electron microscopy.

Biophysical modelling of intra-ring variations in tracheid features and wood density of Pinus pinaster trees exposed to seasonal droughts.

Process-based models that link seasonally varying environmental signals to morphological features within tree rings are essential tools to predict tree growth response and commercially important wood quality traits under future climate scenarios. This study evaluated model portrayal of radial growth and wood anatomy observations within a mature maritime pine (Pinus pinaster (L.) Aït.) stand exposed to seasonal droughts. Intra-annual variations in tracheid anatomy and wood density were identified through image analysis and X-ray densitometry on stem cores covering the growth period 1999-2010. A cambial growth model was integrated with modelled plant water status and sugar availability from the soil-plant-atmosphere transfer model MuSICA to generate estimates of cell number, cell volume, cell mass and wood density on a weekly time step. The model successfully predicted inter-annual variations in cell number, ring width and maximum wood density. The model was also able to predict the occurrence of special anatomical features such as intra-annual density fluctuations (IADFs) in growth rings. Since cell wall thickness remained surprisingly constant within and between growth rings, variations in wood density were primarily the result of variations in lumen diameter, both in the model and anatomical data. In the model, changes in plant water status were identified as the main driver of the IADFs through a direct effect on cell volume. The anatomy data also revealed that a trade-off existed between hydraulic safety and hydraulic efficiency. Although a simplified description of cambial physiology is presented, this integrated modelling approach shows potential value for identifying universal patterns of tree-ring growth and anatomical features over a broad climatic gradient.