Standard Laboratory Protocols Research Articles

English

Ethnobotanical investigation revealed the use of ten medicinal plants in the management of arthritis in Ibadan, Nigeria. This study screened the plants for mineral, proximate and phytochemical components that could be responsible for their therapeutic value in arthritis. The powdered plant samples were analysed for nutritional constituents and phytochemical compounds using standard laboratory protocols. The use value of plant-parts was 50% leaves and 50% roots. Three out of the 10 plants had high calcium content: Oncoba spinosa (180.0 mg/100 g), Nymphaea lotus (160.0 mg/100 g) and Solenostemon monostachyus (125.0 mg/100 g). N. lotus had the highest iron content (8.0 mg/100 g). Phosphorus content was highest in O. spinosa (150.0 mg/100 g). Magnesium was highest in Phyllanthus amarus (14.0 mg/100 g). Crude fibre was highest in Solanum aethiopicum (15.90%) and the least in O. spinosa (14.00%). S. aethiopicum had the highest protein content (18.50%) and O. spinosa the least (14.75%). All the medicinal plants tested positive to alkaloids, carotenoids and flavonoids. The plants contained minerals and secondary metabolites that are implicated in arthritis viz. calcium, zinc, carotenoids and flavonoids. The presence of these compounds in the test plants might alleviate pains associated with arthritis. O. spinosa had high potential in the management of arthritis due to its high calcium and phosphorus components.   Key words: Arthritis, medicinal plants, mineral analysis, proximate analysis, Nigeria.

English

Bacterial vaginosis is a disease affecting many women all over the world, being asymptomatic or symptomatic and producing obstetric and gynaecological complications. The aim of this study was to compare three methods for the diagnosis of bacterial vaginosis. For this cross-sectional descriptive study, vaginal swabs from 101 women attending Máximo Gómez Clinic in Holguín, Cuba, from April to May 2011 were used. One clinical and two laboratory methods were applied. The specimens were processed according to the standard ethical and laboratory protocols. Kappa index was used to determine the concordance between two methods at once, and categories from three methods were compared and analysed. A total of 37 (36.6%), 29 (28.7%) and 42 (41.6%) patients were diagnosed with BV according to Amsel’s, Nugent’s and Claeys’ methods, respectively. Kappa index between methods was 0.6872 (Amsel-Nugent), 0.7522 (Nugent-Claeys) and 0.8134 (Amsel-Claeys). As predictor of BV, from Amsel’s criteria, homogeneous white-greyish discharge was the least specific and whiff test was the most specific, while whiff test was the least sensitive and pH exceeding 4.5 was the most sensitive. The diagnosis of BV by the three methods shows moderate concordance, being Claeys’ method the most effective. Nugent’s method seems to make a sub-diagnosis of the disease and Amsel’s method contains criteria with poor sensitivity and specificity.   Key words: Bacterial vaginosis, polybacterial disease, kappa index, diagnosis, Amsel’s method, Nugent’s method, Claeys’ method

Isolation of MRSA, ESBL and AmpC - β -lactamases from Neonatal Sepsis at a Tertiary Care Hospital.

The emergence of methicillin resistant Staphylococcus aureus (MRSA) and extended spectrum β-lactamases (ESBLs) in neonatal intensive care unit patients is increasing. This study aims to find out the bacteriological profile in neonatal sepsis and study their antimicrobial susceptibility pattern including detection of MRSA and ESBLs. This study was conducted for a period of one and a half years from January 2010 to June 2011 in a tertiary care hospital in Chennai. A total of 182 blood samples were collected using sterile precautions. They were processed following standard laboratory protocol. Antibiogram was done using appropriate antibiotics by Kirby-Bauer disc diffusion method. Isolated Staphylococcus aureus were tested for methicillin resistance using Cefoxitin disc (30μg), ESBL was detected using combined disc method, MIC reduction and Polymerase chain reaction, metallobetalactamases using EDTA and Amp-C beta lactamases using AmpC disc test. C-reactive protein (CRP) was estimated for all the cases. Out of the 182 cases, 110 (60.4%) were culture positive. Fifty five (63.9 %) of early onset sepsis cases had Gram negative bacteria (GNB) and 19 (79.1%) of late onset sepsis cases had Gram positive bacteria. Out of the total pathogens, 31 (28.1%) were Klebsiella pneumoniae and 30 (27%) were Staphylococcus aureus. 17 (56.6 %) of Staphylococcus aureus were found to be MRSA and they were 100% sensitive to Vancomycin. 33 (67.3%) of Enterobacteriaceae were ESBL producers. ESBL isolates were 100% sensitive to Imipenem. Three (6.1%) of Enterobacteriaceae were AmpC producers and 3 (27.2%) of Pseudomonas aeruginosa were MBL producers. CRP was positive in 99 (54.3%) cases, out of which 94 (94.9%) were culture positive. Klebsiella spp. and Staphylococcus aureus were the commonest bacteria causing neonatal sepsis in this centre. Multidrug resistance among the isolates was common. Early diagnosis and institution of specific antibiotics after studying the sensitivity pattern will help in reducing neonatal morbidity and mortality and prevent emergence of drug resistant strains.

Alpha-1-Antitrypsin Deficiency – Molecular Basis, Clinical Presentation, Therapeutic Options and an Integrative Approach in Diagnostics / DEFICIJENCIJA ALFA-1-ANTITRIPSINA – MOLEKULSKE OSNOVE, KLINIČKE MANIFESTACIJE, TERAPIJSKE MOGUĆNOSTI I INTEGRATIVNI PRISTUP U DIJAGNOSTICI

Summary The primary role of alpha-1-antitrypsin (AAT), encoded by the highly polymorphic SERPINA1 gene, is to protect the lung parenchyma from proteolysis by neutrophil elastase. AAT deficiency (AATD) is an autosomal recessive disease, considered as the most important genetic cause of liver disease in children and emphysema in adults. According to frequency, deficient alleles can be classified as »common« (Z and S) and »rare« (Mmalton, Mheerlen, Mprocida etc). Type, intensity and onset of clinical disease associated with AATD occur as a result of interaction between AATD and additional genetic and acquired factors (tobacco smoking, air pollution exposure etc). The most frequent clinical manifestations include premature emphysema, chronic hepatitis, cirrhosis and hepatocellular carcinoma. Epidemiological studies highlight the need for improvement in diagnostic efficiency for AATD. It is recommended for a diagnostic approach to integrate precise, internationally recognized clinical criteria and a standardized laboratory protocol, based on a combination of biochemical and molecular methods. The predilection site of cli nical manifestations guides the therapeutic approach. Augmen tation therapy is possible in lung disease, while currently the only specific measure in patients with severe liver failure due to AATD is transplantation. In all patients, preventive measures, ammeliorating the deleterious effects of habits and environmental factors are recommended. Introduction of gene therapy is expected to additionally improve health outcomes in affected persons. Current results with an integrative AATD diagnostic strategy in the Serbian population are highly encouraging, prompting towards its further implementation in common medical practice with the ultimate goal to establish a national register of affected individuals.

Prognostic Significance Of Cytogenetic Complexity and Del(17p) At Diagnosis Of Chronic Lymphocytic Leukemia (CLL)

Complexity has been associated with an adverse outcome in CLL (Kujawski et al. Blood 2008, 112:1993), and cytogenetic complexity often evolves as the patient progresses (Blood 2013, 121:1403). We sought to determine whether the degree of complexity as determined by cytogenetic analyses correlated with time-to-treatment (TTT) or with overall survival (OS) and its relationship with del(17p).Patients (pts) with CLL who had cytogenetic analyses completed between December 2006 and May 2011 were identified from The Ohio State University Wexner Medical Center (OSU) databases. The first cytogenetic analysis evaluated within one year of diagnosis and prior to treatment was considered the diagnostic sample. All but one cytogenetic analyses were performed using CpG +/- pokeweed mitogen and phorbol 12-myristate 13-acetate stimulation. Cytogenetic methods and analyses were by standard laboratory protocols. Fluorescence in situ hybridization (FISH) was performed on all cases. For these analyses, probes for ATM (11q22.3) and TP53 (17p13.1) (Abbott Molecular, Des Plaines, IL) were used according to the manufacturer's recommendations. Complex karyotype (CK) was defined as ≥3 independent aberrations. Associations between CK and clinical or molecular characteristics were tested using Fisher's exact or Wilcoxon rank sum tests. TTT was calculated from the date of diagnosis until the date of treatment, censoring pts who had not started treatment at the date last seen. OS was calculated from the date of diagnosis until the date of death or last follow-up. Estimates of TTT and OS were calculated using the Kaplan-Meier method, and differences in survival distributions were compared using the log-rank test.Diagnostic samples meeting the above criteria were identified in 143 pts. Median age was 58 (range 33-85), 38% were female, and 44% had Rai stage 0. Twenty pts (14%) had del(11q) and 16 (11%) had del(17p), 10 of which were identified by both FISH and karyotype, 4 by FISH alone, and 2 by karyotype alone. CK was detected in 22 pts (15%, 95% CI: 0.10-0.22). CK tended to be more prevalent in males (20% vs 7% in females, p=0.05) and in those with Rai stage ≥ I (20% vs 8% with Rai stage 0, p=0.06), but there was no significant association with age (p=0.98). Only 5 CK pts (23%) had del(11q) compared to 12% without CK (NCK; p=0.20). CK was strongly associated with del(17p), where 41% of CK pts harbored this typically high-risk aberration versus only 6% in NCK pts (p<0.0001).TTT was significantly shorter in pts with CK compared to NCK (p=0.0005), with 12 mos estimates of 55% (95%CI=0.31-0.74) and 83% (95%CI=0.75-0.89), respectively (Figure 1A). In the subsets of pts with and without del(17p), the negative impact of CK persisted as shown in Figures 2A-B. In a proportional hazards model for TTT, CK remained significantly associated with shorter TTT independent of del(17p) and Rai stage (hazard ratio=2.20, 95%CI=1.03-4.71; p=0.04).With a median follow-up of 32 months for OS, there have been only 14 events. Still, early deaths tended to occur more often in those with CK than NCK; five deaths occurred in the first 12 mos following diagnosis, 3 in the CK group (13.6%) and 2 in the NCK group (1.7%), and hence there was a significant difference in OS (p=0.015; Figure 1B). The limited information prevented meaningful comparisons between CK and NCK by del(17p) subgroups.In summary, CK is indicative of both shorter TTT and OS. Although there is a strong correlation of del(17p) with CK, presence of CK appears to predict TTT independent of del(17p). Complexity should be evaluated in a larger, prospective series to further clarify its role in clinical outcome.▪▪ Disclosures:No relevant conflicts of interest to declare.

Plasma Processing Conditions Substantially Influence Circulating microRNA Biomarker Levels

Circulating, cell-free microRNAs (miRNAs) are promising candidate biomarkers, but optimal conditions for processing blood specimens for miRNA measurement remain to be established. Our previous work showed that the majority of plasma miRNAs are likely blood cell-derived. In the course of profiling lung cancer cases versus healthy controls, we observed a broad increase in circulating miRNA levels in cases compared to controls and that higher miRNA expression correlated with higher platelet and particle counts. We therefore hypothesized that the quantity of residual platelets and microparticles remaining after plasma processing might impact miRNA measurements. To systematically investigate this, we subjected matched plasma from healthy individuals to stepwise processing with differential centrifugation and 0.22 µm filtration and performed miRNA profiling. We found a major effect on circulating miRNAs, with the majority (72%) of detectable miRNAs substantially affected by processing alone. Specifically, 10% of miRNAs showed 4–30x variation, 46% showed 30-1,000x variation, and 15% showed >1,000x variation in expression solely from processing. This was predominantly due to platelet contamination, which persisted despite using standard laboratory protocols. Importantly, we show that platelet contamination in archived samples could largely be eliminated by additional centrifugation, even in frozen samples stored for six years. To minimize confounding effects in microRNA biomarker studies, additional steps to limit platelet contamination for circulating miRNA biomarker studies are necessary. We provide specific practical recommendations to help minimize confounding variation attributable to plasma processing and platelet contamination.

Neuropeptide S receptor gene is associated with cortisol responses to social stress in humans

The neuropeptide S (NPS) and its receptor NPSR represent a transmitter system critically involved in the modulation of anxiety and arousal in rodents. Initial human studies indicate that the T-allele of the functional NPSR gene (NPSR1) polymorphism (rs324981), which increases NPS potency at NPSR, is associated with anxiety-related phenotypes. Since stress is critically involved in the pathogenesis of anxiety disorders, we tested the association between rs324981 and stress reactivity in 196 healthy males. Participants were exposed to the Trier Social Stress Test for Groups (TSST-G), a standardized laboratory protocol for stress exposure in a group format. Salivary cortisol and subjective stress responses were assessed. A significant genotype by time interaction and a main effect of genotype were shown, with T-allele carriers displaying larger cortisol and subjective stress responses. This is the first report to show involvement of the NPS system in the regulation of the neuroendocrine stress response in humans.

Miniscrew design and bone response: Defining a correlation

This prospective clinical trial aims at correlating miniscrew implant (MSI) micro/macro architecture, the method of placement, and biologic markers in peri-MSI crevicular fluid (PMICF) as indicators of bone response. A comparative evaluation of surface morphology of the MSIs before placement and after retrieval defines a correlation between the architecture of the MSIs and the bone- implant contact ratio. Two types of MSIs (hybrid and cylindric) were placed in ten patients using a split-mouth technique with the aid of a restricted random number table. Each of the MSIs was placed in the intraradicular area between the second premolar and first molar in the attached gingiva, 4 mm from the cementoenamel junction. The MSIs were immediately loaded, and PMICF was collected on days 0, 7, 14, 21, 28, and 42 and evaluated using a standard laboratory protocol. Surface morphology before placement and after retrieval of the MSI was observed using scanning electron microscopy (SEM) at a magnification of ×11, ×40, and ×1,000. Alkaline phosphatase (ALP) and aspartate aminotransferase (AST) levels observed were lower in the hybrid MSI in comparison to the cylindric MSI. For both MSIs, ALP and AST levels showed a trend of significant increase at days 0, 7, and 14 and then a significant decrease on days 21, 28, and 42. Observations from SEM showed an oxide layer over the entire surface of the bone-expanding hybrid MSI; this layer was observed only at the tip of the cylindric MSI. Levels of both the diagnostic tissue destruction biologic markers ALP and AST are significantly higher in cylindric MSIs compared with hybrid MSIs, indicating a correlation to the type and method of placement of the MSI. The inflammatory markers show a definitive trend, with an elevation until day 14 and a decline after that, indicating an active inflammatory process until day 14 that could be correlated to tissue trauma. Observations from the SEM show a greater oxide layer formation in the hybrid MSI, which could imply a better bone-MSI contact ratio.

1093P - BCR ABL Fusion Protein Detection by a Modern Approach: Fluroscent Immuno Bead Assay

ABSTRACT Introduction Chronic Myeloid Leukemia (CML) is probably the most extensively studied human malignancy. CML patients (90-95%) harbor the Philadelphia (Ph) chromosome, a shortened chromosome 22, resulting from a reciprocal translocation t (9; 22) (q34; q11) between the long arms of chromosome 9 and 22, fusion ABL proto-oncogene on chromosome 9 with the BCR gene on chromosome 22. BCR-ABL fusion gene plays a key role in pathogenesis of the disease. Approximately 15-30% of adult and 2-5% of childhood ALL and Materials and methods Study Period: June 2009- February 2012 Test Specimen Peripheral blood or Bone marrow from 242 CML patient (from hospital registry of a tertiary cancer institute of Eastern India) Normal (as negative control): Peripheral blood from 20 individuals BCR-ABL positive cell line: K562 FISH was performed using (for 100 samples) directly labeled dual color prob for detection of BCR-ABL as standard laboratory protocol. Isolation of WBC were done by lysis buffer. Then lysate treatment were done to release the intracellular BCR-ABL protein. After fluorescent immune bead assay were performed by captured bead and detector reagent. After forming a sandwich complex (if sample contain targeted protein), they were analysed by flowcytometry method. Result In 242 cases 166 (68.6%) were positive for BCR ABL fusion protein by Flowcytometric Bead Assay. Approximately 96% cases were observed as positive by both techniques FISH and Flowcytometric Bead Assay. Conclusion We conclude that the Flowcytometry Immuno Bead Assay is a faster, easier and reliable technique for detection of BCR ABL protein in leukemic cells and shows promise for serial evaluation of patients undergoing treatment. The main advantage of immune bead assay is not dependence on the breakpoint position in the BCR gene. Moreover this technique is based on protein translation label. Disclosure All authors have declared no conflicts of interest.

Surgical site infections linked to contaminated surgical instruments

This study describes an investigation into a sudden increase in surgical site infection rate following 'clean' surgery. The outbreak involved 15 orthopaedic patients following metal insertion, and five ophthalmology patients who developed endophthalmitis. An outbreak committee was convened in order to find the cause of the sudden increase in surgical infections. The investigation included epidemiological and patient analyses, and environmental and clinical audits of wards and theatres. Following reports of contaminated surgical sets, surgical instruments and their packaging were examined using a standardized laboratory protocol. Clinical staff visited the sterilization plant. Skin flora including coagulase-negative staphylococci (CoNS) and Bacillus spp. were recovered from a range of patient specimens. Eleven patients required further surgical attention. Microbiological processing of surgical packs revealed CoNS and Bacillus spp. from inner packaging as well as from instruments themselves. Inspection of the sterilization plant highlighted inadequate maintenance of autoclave components and poor handling practices by staff. This was compounded by lapses in inspection of surgical sets by theatre staff. Cases terminated following a review of operator training, supervision and staffing at the sterilization plant, in conjunction with formal inspection and reporting of damp/stained sets by theatre staff. Post-sterilization contamination of sets containing surgical instruments was linked with an increased rate of deep surgical site infections in orthopaedic and ophthalmic patients. The investigation demonstrates the importance of close collaboration and co-operation between sterile services providers, managers and clinical staff and offers guidance for reducing the risk of contaminated sterile surgical instruments.

Variations in bacterial pathogens causing early onset neonatal septicaemia, according to birth weights - a 5 year study in a referral hospital of West- Bengal

Bacterial sepsis continues to be a major cause of morbidity and mortality in neonates. The features of sepsis being non- specific, a high index of suspicion can only save a precious life. Pathogens of early onset septicaemia may vary, from one country to another and within a country from one place to the other. This study had been undertaken t o understand the variations in the pathogens of early onset neonatal septicaemia according to birth weights ,their susceptibility patterns to commonly used antibiotics in this age of emerging resistance and to utilise this data for better empirical treatment of this potentially fatal conditions. Blood samples from 255 babies belonging to various birth weights and of both sexes with definitive inclusion and exclusion criteria were processeed according to standard laboratory protocols during a five year period, from February 2005-2010, in a rural referral hospital of West Bengal. Culture positivity was found in 63/255 samples (24.70%), comprising of Klebsiella pneumoniae (31.74%) and methicillin sensitive Staphylococcus aureus (31.74%) as predominant isolates. Male: female ratio was 1.25:1. The susceptibility pattern of isolates has been discussed. It is observed that bacteriology of early onset neonatal septicaemia varies according to birth weights. In very low birth weight babies Eshcherichia coli is the predominant organism and culture positivity is highest (87.5%). Where as culture postivity in low birth weight and normal weight babies with sepsis are 32.89% and 29.26% respectively.In low birth weight babies Staphylococcus aureus and in normal weight babies Klebsiella pneumoniae are the predominant organisms. Birth weight of babies and development of neonatal sepsis are significantly associated x2= 7.174, p&lt;0.03. Male babies are more susceptible to develop neonatal sepsis.For initial therapy of early onset neonatal septicemia, Amikacin is the best choice.DOI: http://dx.doi.org/10.3329/bjmm.v4i2.10823

Amelioration of Experimental Autoimmune Encephalomyelitis in C57BL/6 Mice by Photobiomodulation Induced by 670 nm Light

BackgroundThe approved immunomodulatory agents for the treatment of multiple sclerosis (MS) are only partially effective. It is thought that the combination of immunomodulatory and neuroprotective strategies is necessary to prevent or reverse disease progression. Irradiation with far red/near infrared light, termed photobiomodulation, is a therapeutic approach for inflammatory and neurodegenerative diseases. Data suggests that near-infrared light functions through neuroprotective and anti-inflammatory mechanisms. We sought to investigate the clinical effect of photobiomodulation in the Experimental Autoimmune Encephalomyelitis (EAE) model of multiple sclerosis.Methodology/Principal FindingsThe clinical effect of photobiomodulation induced by 670 nm light was investigated in the C57BL/6 mouse model of EAE. Disease was induced with myelin oligodendrocyte glycoprotein (MOG) according to standard laboratory protocol. Mice received 670 nm light or no light treatment (sham) administered as suppression and treatment protocols. 670 nm light reduced disease severity with both protocols compared to sham treated mice. Disease amelioration was associated with down-regulation of proinflammatory cytokines (interferon-γ, tumor necrosis factor-α) and up-regulation of anti-inflammatory cytokines (IL-4, IL-10) in vitro and in vivo.Conclusion/SignificanceThese studies document the therapeutic potential of photobiomodulation with 670 nm light in the EAE model, in part through modulation of the immune response.

Lethal and pre-lethal effects of a fungal biopesticide contribute to substantial and rapid control of malaria vectors.

Rapidly emerging insecticide resistance is creating an urgent need for new active ingredients to control the adult mosquitoes that vector malaria. Biopesticides based on the spores of entomopathogenic fungi have shown considerable promise by causing very substantial mortality within 7–14 days of exposure. This mortality will generate excellent malaria control if there is a high likelihood that mosquitoes contact fungi early in their adult lives. However, where contact rates are lower, as might result from poor pesticide coverage, some mosquitoes will contact fungi one or more feeding cycles after they acquire malaria, and so risk transmitting malaria before the fungus kills them. Critics have argued that ‘slow acting’ fungal biopesticides are, therefore, incapable of delivering malaria control in real-world contexts. Here, utilizing standard WHO laboratory protocols, we demonstrate effective action of a biopesticide much faster than previously reported. Specifically, we show that transient exposure to clay tiles sprayed with a candidate biopesticide comprising spores of a natural isolate of Beauveria bassiana, could reduce malaria transmission potential to zero within a feeding cycle. The effect resulted from a combination of high mortality and rapid fungal-induced reduction in feeding and flight capacity. Additionally, multiple insecticide-resistant lines from three key African malaria vector species were completely susceptible to fungus. Thus, fungal biopesticides can block transmission on a par with chemical insecticides, and can achieve this where chemical insecticides have little impact. These results support broadening the current vector control paradigm beyond fast-acting chemical toxins.

Routinely Obtained Diagnostic Material as a Source of RNA for Personalized Medicine in Lung Cancer Patients

Therapeutic approaches to lung cancer are evolving, with personalized therapy, based on "molecular analysis" of tumors being developed. Given that approximately 90% of patients will not undergo surgery for their disease, an ability to apply these tests to small samples obtained at the time of initial pathological diagnosis is desirable. Studies in this area have produced variable results, and the minimum area of tumor tissue required for analysis has not been defined. Furthermore, such assays have not been widely applied to cytology specimens, which may be the only source of diagnostic material in many cases. Routinely processed biopsy and cytology specimens were microdissected to enrich for tumor cells, followed by RNA extraction using QIAGEN RNeasy kit and cDNA synthesis/reverse-transcriptase polymerase chain reaction for genes including beta-actin, ERCC-1, and RRM-1, according to standard laboratory protocols. Paired biopsy and resection specimens were similarly analyzed. As little as 1 mm² of tumor tissue, from a 10-μm-thick section, may be used to produce RNA suitable for analysis. RNA of adequate quality and quantity for analysis may be obtained from residual, routinely processed biopsy and cytology specimens. There is good correlation between the result obtained on the tumor biopsy specimen and paired blocks from the surgical resection with respect to clinical decision making. Routinely processed clinical diagnostic samples provide a suitable source of RNA for polymerase chain reaction-based molecular analyses, potentially providing personalized medicine to all lung cancer patients.

Integration of 3‐D cell cultures in fluidic microsystems for biological screenings

AbstractA life support system for the cultivation of adherent 2‐D and scaffold‐based 3‐D cell cultures in a microfluidic device, a Bio‐Micro‐Electro‐Mechanical System (BioMEMS) is presented. The miniaturization level and system set‐up allow incubator‐independent operation modes and long‐term experiments with real‐time microscope observation. A dedicated seeding procedure for adherent cells into the microstructures is one key issue of the BioMEMS developed. Several seeding methods for the cells were evaluated. Biocompatibility of all materials, surfaces and methods could be demonstrated. First experiments with several cell types show the feasibility of the approach employing standard laboratory protocols. At present, the modular design and set‐up offer a broad application spectrum as well as its future extension to e.g. cultivation of other cell types, coupled cultivation chambers and the implementation of other manipulation or analysis components.

Social Stratification in the Sikh Population of Punjab (India) Has a Genetic Basis: Evidence from Serological and Biochemical Markers

The present study was planned to assess whether social stratification in the Sikh population inhabiting the northwest border Indian state of Punjab has any genetic basis. Blood samples were collected randomly from a total of 2851 unrelated subjects belonging to 21 groups of two low-ranking Sikh scheduled caste populations, viz. Mazhabi and Ramdasi, and a high-ranking Jat Sikh caste population of Punjab. The genetic profile of Sikh groups was investigated using a total of nine serobiochemical genetic markers, comprising two blood groups (ABO, RH(D)) and a battery of seven red cell enzyme polymorphisms (ADA, AK1, ESD, PGM1, GLO1, ACP1, GPI), following standard serological and biochemical laboratory protocols. Genetic structure was studied using original allele frequency data and statistical measures of heterozygosity, genic differentiation, genetic distance, and genetic admixture. Great heterogeneity was observed between Sikh scheduled caste and Jat Sikh populations, especially in the RH(D) blood group system, and distribution of ESD, ACP1, and PGM1 enzyme markers was also found to be significantly different between many of their groups. Genetic distance trees demonstrated little or no genetic affinities between Sikh scheduled caste and Jat Sikh populations; the Mazhabi and Ramdasi also showed little genetic relationship. Genetic admixture analysis suggested a higher element of autochthonous tribal extraction in the Ramdasi. The present study revealed much genetic heterogeneity in differently ranking Sikh caste populations of Punjab, mainly attributable to their different ethnic backgrounds, and provided a genetic basis to social stratification present in this religious community of Punjab, India.

Determining CO2 /brine relative permeability and capillary threshold pressures for reservoir rocks and caprocks: Recommendations for development of standard laboratory protocols

Important parameters for the determination of fluid transport in reservoir rocks are capillary pressures and relative permeabilities for the CO2/brine system. These factors influence the plume migration, residual CO2 trapping and CO2 dissolution in the reservoir brines. CO2-brine relative permeabilities for different rock types are rare in public literature and usually adapted from mathematical relations or from more recent experimental data. The experimental data vary greatly in saturation and relative permeability endpoints, between 10 and 100%. It is necessary to reduce the uncertainty in the data range, since these parameters are sensitive and indispensable for the numerical simulation of the evolution of a CO2 plume in the subsurface. The petrophysical properties of caprocks are the limiting factor to the amount of CO2 stored in potential storage sites. The caprock sealing capacity is determined by the capillary pressure (or column height) at which the trapped fluid starts leaking through a caprock. These rocks have high capillary threshold pressures, which, combined with wettability and interfacial tension (IFT), determine the final column height that a seal can hold. Those three factors - capillary pressure, wettability and IFT - play an important role in the geological storage of CO2 in controlling the potential of CO2 moving through the seal. The capillary threshold pressures need to be determined accurately toestimate brine displacement in caprocks and hence the onset of viscous flow or leakage to the surface. Within the framework of the Carbon Capture Project (CCP3), we plan to establish standard experimental protocols with academic and/or commercial laboratories.

Sampling for Pharmaceuticals and Personal Care Products (PPCPs) and Illicit Drugs in Wastewater Systems: Are Your Conclusions Valid? A Critical Review

The analysis of 87 peer-reviewed journal articles reveals that sampling for pharmaceuticals and personal care products (PPCPs) and illicit drugs in sewers and sewage treatment plant influents is mostly carried out according to existing tradition or standard laboratory protocols. Less than 5% of all studies explicitly consider internationally acknowledged guidelines or methods for the experimental design of monitoring campaigns. In the absence of a proper analysis of the system under investigation, the importance of short-term pollutant variations was typically not addressed. Therefore, due to relatively long sampling intervals, potentially inadequate sampling modes, or insufficient documentation, it remains unclear for the majority of reviewed studies whether observed variations can be attributed to "real" variations or if they simply reflect sampling artifacts. Based on results from previous and current work, the present paper demonstrates that sampling errors can lead to overinterpretation of measured data and ultimately, wrong conclusions. Depending on catchment size, sewer type, sampling setup, substance of interest, and accuracy of analytical method, avoidable sampling artifacts can range from "not significant" to "100% or more" for different compounds even within the same study. However, in most situations sampling errors can be reduced greatly, and sampling biases can be eliminated completely, by choosing an appropriate sampling mode and frequency. This is crucial, because proper sampling will help to maximize the value of measured data for the experimental assessment of the fate of PPCPs as well as for the formulation and validation of mathematical models. The trend from reporting presence or absence of a compound in "clean" water samples toward the quantification of PPCPs in raw wastewater requires not only sophisticated analytical methods but also adapted sampling methods. With increasing accuracy of chemical analyses, inappropriate sampling increasingly represents the major source of inaccuracy. A condensed step-by-step Sampling Guide is proposed as a starting point for future studies.

Study of Framing Effects in a New Risk Aversion Experiment

Risk aversion experiments such as those by Holt and Laury (2002 and 2005) measure risk aversion by examining responses of experimental subjects who are confronted with single-sheet paper displays of probability-ordered arrays of choices in which “real” money is at risk. As an alternative to this approach, the findings reported in this paper were obtained using a modified adventure-type video game to offer the choices presented by the HL experiment embedded in a more realistic scenario. The decisions are confronted first by our experimental subjects in a sequential and unordered manner. Then, later in the experiment, subjects are instructed to examine the results of their decisions in an array that shares the simultaneous and probability-ordered characteristics of the standard laboratory protocol. Subjects then had the option of altering their decisions before their payment was determined. The results indicate that decisions made in a sequential and unordered manner exhibit less risk aversion and higher degrees of decision inconsistency.

Stathmin, a gene regulating neural plasticity, affects fear and anxiety processing in humans

The identification of biological mechanisms underlying emotional behavior is crucial for our understanding of the pathogenesis of mental disorders. Besides genes modulating neural transmission and influencing amygdala reactivity and anxiety-related temperamental traits a different plasticity regulating genes affect interindividual differences in emotional regulation. Recently it has been demonstrated that stathmin, a regulator of microtubule formation which affects long-term potentiation (LTP), controls learned and innate fear responses in rodents, but its role in human emotion regulation is unknown. We hypothesized that in humans the gene coding for stathmin (STMN1), which is highly expressed in the lateral nucleus of the amygdala and associated thalamic and cortical structures, influences behavioral responses to fear and anxiety stimuli by way of two common single nucleotide polymorphisms (rs182455, SNP1; rs213641, SNP2). These polymorphisms are located within or close to the putative transcriptional control region. We used the acoustic startle paradigm and a standardized laboratory protocol for the induction of fear and psychosocial stress in 106 healthy volunteers to investigate the impact of stathmin gene variation on two fear- and anxiety-controlling effector-systems of the amygdala. We found that STMN1 genotype interacting with individuals' gender significantly impacts fear and anxiety responses as measured with the startle and cortisol stress response. We therefore conclude that STMN1 genotype has functional relevance for the acquisition and expression of basic fear and anxiety responses also in humans.