DPPrimer – A Degenerate PCR Primer Design Tool

Designed degenerate primers unlike conventional primers are superior in matching and amplification of large number of genes, from related gene families. DPPrimer tool was designed to predict primers for PCR amplification of homologous gene from related or diverse plant species. The key features of this tool include platform independence and user friendliness in primer design. Embedded features such as search for functional domains, similarity score selection and phylogebetic tree further enhance the user friendliness of DPPrimer tool. Performance of DPPrimer tool was evaluated by successful PCR amplification of ADP-glucose phosphorylase genes from wheat, barley and rice.

Availability

DPPrimer is freely accessible at http://202.141.12.147/DGEN_tool/index.html     

Introduction to Biostatistics—An Annotated Bibliography for Medical Researchers

Biostatistical methods have become thoroughly integrated into modern biomedical and clinical research. Nevertheless, every observer who has evaluated articles in medical journals has noted that as many as half the reported results were based on questionable statistical analysis. This situation, combined with the fact that most errors involve relatively simple statistical procedures, points to the need for researchers and practitioners to be able to personally judge the quality of the statistical analyses in what they read. Fortunately, there are several excellent papers and texts available for those interested.     

Quantitative Assessment of Orbital Implant Position – A Proof of Concept

Introduction

In orbital reconstruction, the optimal location of a predefined implant can be planned preoperatively. Surgical results can be assessed intraoperatively or postoperatively. A novel method for quantifying orbital implant position is introduced. The method measures predictability of implant placement: transformation parameters between planned and resulting implant position are quantified.

Methods

The method was tested on 3 human specimen heads. Computed Tomography scans were acquired at baseline with intact orbits (t0), after creation of the defect (t1) and postoperatively after reconstruction of the defect using a preformed implant (t2). Prior to reconstruction, the optimal implant position was planned on the t0 and t1 scans. Postoperatively, the planned and realized implant position were compared. The t0 and t2 scans were fused using iPlan software and the resulting implant was segmented in the fused t2 scan. An implant reference frame was created (Orbital Implant Positioning Frame); the planned implant was transformed to the reference position using an Iterative Closest Point approach. The segmentation of the resulting implant was also registered on the reference position, yielding rotational (pitch, yaw, roll) as well as translational parameters of implant position.

Results

Measurement with the Orbital Implant Positioning Frame proved feasible on all three specimen. The positional outcome provided more thorough and accurate insight in resulting implant position than could be gathered from distance measurements alone. Observer-related errors were abolished from the process, since the method is largely automatic.

Conclusion

A novel method of quantifying surgical outcome in orbital reconstructive surgery was presented. The presented Orbital Implant Positioning Frame assessed all parameters involved in implant displacement. The method proved to be viable on three human specimen heads. Clinically, the method could provide direct feedback intraoperatively and could improve postoperative evaluation of orbital reconstructive surgery.     

A Novel Quantitative Approach to Women’s Reproductive Strategies

The patterned way in which individuals allocate finite resources to various components of reproduction (e.g. mating effort, reproductive timing and parental investment) is described as a reproductive strategy. As energy is limited, trade-offs between and within aspects of reproductive strategies are expected. The first aim of this study was to derive aspects of reproductive strategies using complete reproductive histories from 718 parous Western Australian women. Factor analysis using a subset of these participants resulted in six factors that represented ‘short-term mating strategy’, ‘early onset of sexual activity’, ‘reproductive output’, ‘timing of childbearing’, ‘breastfeeding’, and ‘child spacing’. This factor structure was internally validated by replication using a second independent subset of the data. The second aim of this study examined trade-offs between aspects of reproductive strategies derived from aim one. Factor scores calculated for each woman were incorporated in generalised linear models and interaction terms were employed to examine the effect of mating behaviour on the relationships between reproductive timing, parental investment and overall reproductive success. Early sexual activity correlates with early reproductive onset for women displaying more long-term mating strategies. Women with more short-term mating strategies exhibit a trade-off between child quantity and child quality not observed in women with a long-term mating strategy. However, women with a short-term mating strategy who delay reproductive timing exhibit levels of parental investment (measured as breastfeeding duration per child) similar to that of women with long-term mating strategies. Reproductive delay has fitness costs (fewer births) for women displaying more short-term mating strategies. We provide empirical evidence that reproductive histories of contemporary women reflect aspects of reproductive strategies, and associations between these strategic elements, as predicted from life history theory.     

Risk Factors for Hospitalization Due to Community-Acquired Sepsis – A Population-Based Case-Control Study

Background

The aim of the study was to estimate risk factors for hospitalization due to sepsis and to determine whether these risk factors vary by age and gender.

Methods

We performed a population-based case-control study of all adult patients admitted to a medical ED from September 2010 to August 2011. Controls were sampled within the hospital catchment-area. All potential cases were manually validated using a structured protocol. Vital signs and laboratory values measured at arrival were registered to define systemic inflammatory response syndrome and organ dysfunction. Multivariable logistic regression was used to elucidate which predefined risk factors were associated with an increased or decreased risk hospitalization due to sepsis.

Results

A total of 1713 patients were admitted with sepsis of any severity. The median age was 72 years (interquartile range: 57–81 years) and 793 (46.3%) were male. 621 (36.3%) patients were admitted with sepsis, 1071 (62.5%) with severe sepsis and 21 (1.2%) with septic shock. Episodes with sepsis of any severity were associated with older age (85+ years adjusted OR 6.02 [95%CI: 5.09–7.12]), immunosuppression (4.41 [3.83–5.09]), alcoholism-related conditions (2.90 [2.41–3.50]), and certain comorbidities: psychotic disorder (1.90 [1.58–2.27]), neurological (1.98 [1.73–2.26]), respiratory (3.58 [3.16–4.06]), cardiovascular (1.62 [1.41–1.85]), diabetes (1.82 [1.57–2.12]), cancer (1.44 [1.22–1.68]), gastrointestinal (1.71 [1.44–2.05]) and renal (1.46 [1.13–1.89]). The strength of the observed associations for comorbid factors was strongest among younger individuals.

Conclusions

Hospitalization due to sepsis of any severity was associated with several independent risk factors, including age and comorbid factors.     

Getting More Out of Less – A Quantitative Serological Screening Tool for Simultaneous Detection of Multiple Influenza A Hemagglutinin-Types in Chickens

Current avian influenza surveillance in poultry primarily targets subtypes of interest for the veterinary sector (H5, H7). However, as virological and serological evidence suggest, surveillance of additional subtypes is important for public health as well as for the poultry industry. Therefore, we developed a protein microarray enabling simultaneous identification of antibodies directed against different HA-types of influenza A viruses in chickens. The assay successfully discriminated negative from experimentally and naturally infected, seropositive chickens. Sensitivity and specificity depended on the cut-off level used but ranged from 84.4% to 100% and 100%, respectively, for a cut off level of ≥1∶40, showing minimal cross reactivity. As this testing platform is also validated for the use in humans, it constitutes a surveillance tool that can be applied in human-animal interface studies.     

A New Method for Quantitative Immunoblotting of Endogenous α-Synuclein

β-Sheet-rich aggregates of α-synuclein (αSyn) are the hallmark neuropathology of Parkinson’s disease and related synucleinopathies, whereas the principal native structure of αSyn in healthy cells - unfolded monomer or α-helically folded oligomer - is under debate. Our recent crosslinking analysis of αSyn in intact cells showed that a large portion of endogenous αSyn can be trapped as oligomers, most notably as apparent tetramers. One challenge in such studies is accurately quantifying αSyn Western blot signals among samples, as crosslinked αSyn trends toward increased immunoreactivity. Here, we analyzed this phenomenon in detail and found that treatment with the reducible amine-reactive crosslinker DSP strongly increased αSyn immunoreactivity even after cleavage with the reducing agent β-mercaptoethanol. The effect was observed with all αSyn antibodies tested and in all sample types from human brain homogenates to untransfected neuroblastoma cells, permitting easy detection of endogenous αSyn in the latter, which had long been considered impossible. Coomassie staining of blots before and after several hours of washing revealed complete retention of αSyn after DSP/β-mercaptoethanol treatment, in contrast to a marked loss of αSyn without this treatment. The treatment also enhanced immunodetection of the homologs β- and γ-synuclein and of histones, another group of small, lysine-rich proteins. We conclude that by neutralizing positive charges and increasing protein hydrophobicity, amine crosslinker treatment promotes adhesion of αSyn to blotting membranes. These data help explain the recent report of fixing αSyn blots with paraformaldehyde after transfer, which we find produces similar but weaker effects. DSP/β-mercaptoethanol treatment of Western blots should be particularly useful to quantify low-abundance αSyn forms such as extracellular and post-translationally modified αSyn and splice variants.     

Quantitative analysis of anthropogenic influences on coastal water – A new perspective

Coastal environment has been disturbed by human activities for a long time, especially in the rapidly urbanizing and industrializing areas. Although the surrounding area has achieved great economic success in the past 30 years, Western Xiamen Bay (China) is seriously affected by pollutants and is facing increasing ecological pressure. Because of this, Xiamen was selected in 1994 as a demonstration site for implementing an integrated coastal management program, which included a series of measures for protecting the coastal environment. However, coastal environment is dynamic, complex and site-specific, and thus a scientific quantitative evaluation framework is necessary for environment quality analysis and effective coastal management. In this study, we used oceanographic knowledge together with quantitative methods (Bai-Perron's structural break test) to analyze the long-term variations of water quality indices (pH, DO, COD, DIN, PO₄-P and Oil) in Western Xiamen Bay. In addition, we compared with other coastal areas to identify the effectiveness of phosphorus-based nutrient management measures and predicted the probable variation trend in the future. The results show that in Western Xiamen Bay: (1) the concentrations of DO and Oil in seawater are effectively controlled by local coastal management measures; (2) seawater acidification will continue to worsen based on the present situation; and (3) the P-limitation treatment strategies are effective and PO₄-P concentration starts to fall according to the multiple statistical analysis and Environmental Kuznets Curve. This paper hopes to provide an epitome of the conflicts and consolations between socioeconomic development and environmental quality in the past, and hints for coastal management in the future.     

Arbitrary Primer Based RAPD — A Useful Genetic Marker for Species Identification in Morels

Internal transcribed spacer (ITS) regions of 5.8S ribosomal DNA gene of 46 monospore cultures representing different species of family Morchellaceae were sequenced. Eight putative species namely Morchella esculenta, M. crassipes, M. angusticeps, M. conice, Mitrophora semilibera, Morchella spongiola, M. vulgaris and Verpa conica were designated to the sequenced cultures analyzed based on similarity with the best aligned sequence of the BLAST search. We recorded interspecific polymorphism in ITS regions of the 5.8S r-DNA gene amongst putative species of morels. However no intraspecific ITS polymorphism could be visualized amongst monospores from single and or different ascocarps collected from various geographical regions. PCR RAPDamplified profiles of different monospores were identical at intraspecific levels and could not be distinguished by eight arbitrary primers tested. RAPD profiles exhibited significant interspecific polymorphism distinguishing all the eight putative species from each other by primer OPP-6 (5’- GTG GGT TGA C- 3’) and a custom primer (5’-CGC ACC GCA G-3’). RAPDprofiles generated using aforesaid arbitrary primers can serve as useful genetic markers for species identification and to improve morel systematics. Present study is the first report on exploitation of random primers in species differentiation in morels.     

Whakapapa – A Foundation for Genetic Research?

Whakapapa is the foundation of traditional Māori social structure and it perpetuates a value base that locates people through their relationships to the physical and spiritual worlds. As part of a new envirogenomics research programme, researchers at the Institute of Environmental Science and Research (ESR) are developing a study with an iwi (tribe) to identify combinations of genetic and environmental factors that may contribute to current health status. A major objective of this study is to utilise whakapapa (genealogical information) to explore patterns of genetic variation unique to the iwi and to correlate these with potential disease or ill health. Genetic testing and screening raises numerous ethical issues, particularly when indigenous peoples are the subjects. This paper will outline ESR’s strategy for addressing indigenous concerns about genetic testing and how whakapapa forms an integral part of the envirogenomics research programme.     

Dispersal-limited species – A challenge for ecological restoration

In recent years, increasing efforts have been directed to the restoration of natural communities affected by human activities. However, restoration success is often reduced because of the lack of dispersal-limited species in the restored site. Numerous rare species are dispersal-limited, having low probabilities to re-colonize restored sites and to establish viable populations. A high species richness enhances the ability of ecosystems to maintain multiple functions. It has been shown that in highly diverse ecosystems such as alpine meadows, tropical forests and coral reefs, the most distinct combinations of functions are supported predominantly by rare species. Thus, more emphasis should be given to support rare species in restoration actions. I suggest that well-known techniques for increasing plant diversity in restoration projects should be complemented with techniques to re-introduce dispersal-limited animals. Re-establishing ecological connectivity might be of benefit for certain species in the long term. However, for the majority of dispersal-limited animal species, translocations of founder individuals might be the only tool to re-establish populations in restored sites.     

Antibiotic adjuvants – A strategy to unlock bacterial resistance to antibiotics

Resistance to available antibiotics in pathogenic bacteria is currently a global challenge since the number of strains that are resistant to multiple types of antibiotics has increased dramatically each year and has spread worldwide. To unlock this problem, the use of an ‘antibiotic adjuvant’ in combination with an antibiotic is now being exploited. This approach enables us to prolong the lifespan of these life-saving drugs. This digests review provides an overview of the main types of antibiotic adjuvants, the basis of their operation and the remaining issues to be tackled in this field. Particular emphasis is placed on those compounds that are already in clinical development, namely β-lactamase inhibitors.     

A Quantitative Model for Nonrandom Generalized Transduction, Applied to the Phage P22–SALMONELLA TYPHIMURIUM System

A mathematical model for nonrandom generalized transduction is proposed and analyzed. The model takes into account the finite number of transducing particle classes for any given marker. The equations for estimation of the distance between markers from contransduction frequency data are derived and standard errors of the estimates are given. The obtained relationships depend significantly on the number of classes of transducing fragments. The model was applied to estimate the number of transducing fragment classes for a given marker in transduction with phage P22 of Salmonella typhimurium. It was found that the literature data on frequencies of contransduction in crosses with mutual substitution of selective and nonselective markers can be rationalized most accurately by assuming that the mean number of classes is equal to 2. An improved method for analysis of cotransduction data is proposed on the basis of our model and the results of calculation. The method relies on solving a set of algebraic equations for cotransduction frequencies of markers located within one phage length. The method allows a relatively precise determination of distances between markers, positions of transducing particle ends and deletion or insertion lengths. The approach is applied to the trp-cysB-pyrF and aroC-hisT-purF-dhuA regions of the Salmonella typhimurium chromosome.     

Pre-Darwinian Taxonomy and Essentialism – A Reply to MaryWinsor

Mary Winsor (2003) argues against the received view that pre-Darwinian taxonomy was characterized mainly by essentialism. She argues, instead, that the methods of pre-Darwinian taxonomists, in spite of whatever their beliefs, were that of clusterists, so that the received view, propagated mainly by certain modern biologists and philosophers of biology, should at last be put to rest as a myth. I argue that shes right when it comes to higher taxa, but wrong when it comes the most important category of all, the species category.