Real-time PCR quantitation of hepatitis B virus DNA using automated sample preparation and murine cytomegalovirus internal control

Quantitation of circulating hepatitis B virus (HBV) DNA is important for monitoring disease progression and for assessing the response to antiviral therapy. Several commercial and 'in house' assays for HBV DNA quantitation have been described but many of these have limitations of relatively low sensitivity and limited dynamic range. This study describes the development and evaluation of a FRET-based real-time PCR assay designed to overcome these limitations and to provide accurate quantitation of DNA from all eight genotypes of HBV (A-H). The assay employs a fully automated nucleic acid extraction system permitting high-sample throughput with minimal 'hands-on' time and incorporates a murine cytomegalovirus (mCMV) internal control to prevent false negative results and under-reporting due to unrecognised problems with viral lysis, DNA purification or PCR amplification. Sensitivity, assessed by Probit analysis at the 95% detection level, was 24.4 IU/ml, associated with an extremely wide dynamic range (similar to 9 log(10)). Coefficients of variation were low for both intra-assay and inter-assay variability (CV%, 7-11%) and quantitative data correlated well (R-2 = 0.97) with the Digene hybrid capture assay. This assay provides an ideal system for therapeutic monitoring and for studying the relationship between HBV viral load and stage of disease. (c) 2005 Elsevier B.V. All rights reserved

Entrepreneurial capital, social values and Islamic traditions: exploring the growth of women-owned enterprises in Pakistan

Main ArticleThis study seeks to explore the variables contributing to the growth of women-owned enterprises in the Islamic Republic of Pakistan. Based on a previously established multivariate model, it uses two econometric approaches: first classifying variables into predetermined blocks; and second, using the general to specific approach. Statistical analyses and in-depth interviews confirm that women entrepreneurs’ personal resources and social capital have a significant role in their business growth. Further, it reveals that the moral support of immediate family, independent mobility and being allowed to meet with men play a decisive role in the sales and employment growth of women-owned enterprises in an Islamic country such as Pakistan

The analysis of acetaminophen (paracetamol) and seven metabolites in rat, pig and human plasma by U(H)PLC–MS

A U(H)PLC–MS/MS method is described for the analysis of acetaminophen and its sulphate, glucuronide, glutathione, cysteinyl and N-acetylcysteinyl metabolites in plasma using stable isotope-labeled internal standards. P-Aminophenol glucuronide and 3-methoxyacetaminophen were monitored and semi-quantified using external standards. The assay takes 7.5 min/sample, requires only 5 μl of plasma and involves minimal sample preparation. The method was validated for rat plasma and cross validated for human and pig plasma and mouse serum. LOQ in plasma for these analytes were 0.44 μg/ml (APAP-C), 0.58 μg/ml (APAP-SG), 0.84 μg/ml (APAP-NAC), 2.75 μg/ml (APAP-S), 3.00 μg/ml (APAP-G) and 16 μg/ml (APAP). Application of the method is illustrated by the analysis of plasma following oral administration of APAP to male Han Wistar rats

24-hour efficacy of once-daily desloratadine therapy in patients with seasonal allergic rhinitis [ISRCTN32042139]

BACKGROUND: Early studies with desloratadine demonstrated efficacy in treating seasonal allergic rhinitis (SAR). A dose-ranging study was conducted to characterize its 24-hour efficacy in patients with SAR. METHODS: Patients (N = 1,026) were randomly assigned once-daily (QD) desloratadine (2.5, 5, 7.5, 10, or 20 mg) for 2 weeks in a placebo-controlled, double-blind study. The end point of 24-hour efficacy was assessed by the mean change from baseline in the average AM instantaneous total symptom score (TSS) over the treatment period. Day 2 data were assessed for efficacy of desloratadine following the first dose. Other efficacy variables included AM/PM previous total nasal and nonnasal symptom scores and individual symptom scores. RESULTS: Desloratadine 5–20 mg was significantly (P < .01) more effective than placebo in improving total AM instantaneous TSS and AM/PM previous total nasal and nonnasal symptom scores. This dosing range also was significantly (P < .01) more effective than placebo for reducing AM instantaneous TSS beginning with the first dose; thus, demonstrating the full 24-hour efficacy of desloratadine. AM/PM previous scores for all individual symptoms, including nasal congestion, were also significantly improved versus placebo (P < .05) with desloratadine at 5, 7.5, and 20 mg. All treatments were well tolerated. There were no clinically meaningful changes in electrocardiogram parameters. CONCLUSION: Desloratadine 5–20 mg provided significant 24-hour relief of SAR signs and symptoms. There were no statistically significant differences between the 4 largest doses suggesting that desloratadine 5 mg QD offers the best therapeutic profile for patients with SAR

Reservoir Engineering of Wairakei Geothermal Field

Wairakei was the first liquid dominated geothermal field exploited for major power production. As such many decisions were taken on an ad-hoc or experimental basis. In retrospect the choice of Wairakei was fortunate : with extensive shallow high permeability and major recharge it is an easy field to exploit. This lecture describes the history of the field and the contribution of reservoir engineering to field management, and describes the reservoir as it is now understood

Evaluating the potential for the environmentally sustainable control of foot and mouth disease in Sub-Saharan Africa

Strategies to control transboundary diseases have in the past generated unintended negative consequences for both the environment and local human populations. Integrating perspectives from across disciplines, including livestock, veterinary and conservation sectors, is necessary for identifying disease control strategies that optimise environmental goods and services at the wildlife-livestock interface. Prompted by the recent development of a global strategy for the control and elimination of foot-and-mouth disease (FMD), this paper seeks insight into the consequences of, and rational options for potential FMD control measures in relation to environmental, conservation and human poverty considerations in Africa. We suggest a more environmentally nuanced process of FMD control that safe-guards the integrity of wild populations and the ecosystem dynamics on which human livelihoods depend while simultaneously improving socio-economic conditions of rural people. In particular, we outline five major issues that need to be considered: 1) improved understanding of the different FMD viral strains and how they circulate between domestic and wildlife populations; 2) an appreciation for the economic value of wildlife for many African countries whose presence might preclude the country from ever achieving an FMD-free status; 3) exploring ways in which livestock production can be improved without compromising wildlife such as implementing commodity-based trading schemes; 4) introducing a participatory approach involving local farmers and the national veterinary services in the control of FMD; and 5) finally the possibility that transfrontier conservation might offer new hope of integrating decision-making at the wildlife-livestock interface

Genetic Tests for Ecological and Allopatric Speciation in Anoles on an Island Archipelago

From Darwin's study of the Galapagos and Wallace's study of Indonesia, islands have played an important role in evolutionary investigations, and radiations within archipelagos are readily interpreted as supporting the conventional view of allopatric speciation. Even during the ongoing paradigm shift towards other modes of speciation, island radiations, such as the Lesser Antillean anoles, are thought to exemplify this process. Geological and molecular phylogenetic evidence show that, in this archipelago, Martinique anoles provide several examples of secondary contact of island species. Four precursor island species, with up to 8 mybp divergence, met when their islands coalesced to form the current island of Martinique. Moreover, adjacent anole populations also show marked adaptation to distinct habitat zonation, allowing both allopatric and ecological speciation to be tested in this system. We take advantage of this opportunity of replicated island coalescence and independent ecological adaptation to carry out an extensive population genetic study of hypervariable neutral nuclear markers to show that even after these very substantial periods of spatial isolation these putative allospecies show less reproductive isolation than conspecific populations in adjacent habitats in all three cases of subsequent island coalescence. The degree of genetic interchange shows that while there is always a significant genetic signature of past allopatry, and this may be quite strong if the selection regime allows, there is no case of complete allopatric speciation, in spite of the strong primae facie case for it. Importantly there is greater genetic isolation across the xeric/rainforest ecotone than is associated with any secondary contact. This rejects the development of reproductive isolation in allopatric divergence, but supports the potential for ecological speciation, even though full speciation has not been achieved in this case. It also explains the paucity of anole species in the Lesser Antilles compared to the Greater Antilles

BLM and RMI1 alleviate RPA inhibition of topoIIIα decatenase activity

RPA is a single-stranded DNA binding protein that physically associates with the BLM complex. RPA stimulates BLM helicase activity as well as the double Holliday junction dissolution activity of the BLM-topoisomerase IIIα complex. We investigated the effect of RPA on the ssDNA decatenase activity of topoisomerase IIIα. We found that RPA and other ssDNA binding proteins inhibit decatenation by topoisomerase IIIα. Complex formation between BLM, TopoIIIα, and RMI1 ablates inhibition of decatenation by ssDNA binding proteins. Together, these data indicate that inhibition by RPA does not involve species-specific interactions between RPA and BLM-TopoIIIα-RMI1, which contrasts with RPA modulation of double Holliday junction dissolution. We propose that topoisomerase IIIα and RPA compete to bind to single-stranded regions of catenanes. Interactions with BLM and RMI1 enhance toposiomerase IIIα activity, promoting decatenation in the presence of RPA

The genetic contribution of single male immigrants to small, inbred populations: A laboratory study using drosophila melanogaster

This study examined the genetic contribution of single male immigrants to small, inbred laboratory populations of Drosophila melanogaster. Genetic contribution was assessed by measuring the relative frequency of immigrant marker alleles in the first and second generations after immigration, while controlling for any selection effects at the marker locus, and for the experience of male immigrants. When immigrants were outbred, the mean frequency of the immigrant allele was significantly higher than its initial frequency, in both the first and second generations after immigration. There was no significant change in allele frequency for populations receiving inbred immigrants. The increase in allele frequency for outbred immigrants was attributed to an initial outbred vigour fitness advantage of immigrant males over resident males experiencing inbreeding depression. Hybrid vigour of immigrant progeny and the rare-male effect did not have a statistically significant role in the fitness advantage of the immigrant allele. The results suggest that inbreeding may have a considerable impact on the contribution of immigrants to the genetic diversity of populations