Hematocrit and Hemoglobin Determination in the Mangrove Killifish, Kryptolebias marmoratus

Hematocrit, hemoglobin concentration, and mean cell hemoglobin concentration (MCHC) were determined in the mangrove killifish, Kryptolebias marmoratus, a tropical fish that can undergo emersion for \u3e1 month when they leave the waters of mangrove forests and find refuge in moist, terrestrial habitats. Hematocrit was 27.67% ± 1.22 (mean ± s.e.m.), hemoglobin concentration was 7.41 ± 0.52 g/dL, and MCHC was 26.47 ± 0.93 g/dL. A linear relationship was present between hematocrit and hemoglobin in K. marmoratus (R2 = 0.833; P \u3c 0.001) . A significant correlation between collected blood volume and standard length was present (P = 0.001). Limitations of the assays used to measure hemoglobin and hematocrit in this small fish species are discussed. Hematocrit and hemoglobin concentrations are within ranges determined for other air-breathing fishes and show the feasibility of measuring hematological parameters in this small fish during emersion

An Australian childhood obesity summit: the role of data and evidence in 'public' policy making

BACKGROUND: Overweight and obesity in Australia has risen at an alarming rate over the last 20 years as in other industrialised countries around the world, yet the policy response, locally and globally, has been limited. Using a childhood obesity summit held in Australia in 2002 as a case study, this paper examines how evidence was used in setting the agenda, influencing the Summit debate and shaping the policy responses which emerged. The study used multiple methods of data collection including documentary analysis, key informant interviews, a focus group discussion and media analysis. The resulting data were content analysed to examine the types of evidence used in the Summit and how the state of the evidence base contributed to policy-making. RESULTS: Empirical research evidence concerning the magnitude of the problem was widely reported and largely uncontested in the media and in the Summit debates. In contrast, the evidence base for action was mostly opinion and ideas as empirical data was lacking. Opinions and ideas were generally found to be an acceptable basis for agreeing policy action coupled with thorough evaluation. However, the analysis revealed that the evidence was fiercely contested around food advertising to children and action agreed was therefore limited. CONCLUSION: The Summit demonstrated that policy action will move forward in the absence of strong research evidence. Where powerful and competing groups contest possible policy options, however, the evidence base required for action needs to be substantial. As with tobacco control, obesity control efforts are likely to face ongoing challenges around the nature of the evidence and interventions proposed to tackle the problem. Overcoming the challenges in controlling obesity will be more likely if researchers and public health advocates enhance their understanding of the policy process, including the role different types of evidence can play in influencing public debate and policy decisions, the interests and tactics of the different stakeholders involved and the part that can be played by time-limited yet high profile events such as Summits

Community composition of zooplankton exported from a shallow polymictic reservoir linked to wind conditions

Zooplankton exported from lentic systems provision lotic systems with easily captured, consumed, and assimilated prey items. Previous studies have demonstrated that the community composition of zooplankton exports (CCZE) vary over time, which introduces temporal differences in lotic resource availability (zooplankton prey) in downstream habitats. In the study presented here, we monitored variation in CCZE from a polymictic reservoir outfall in response to physical–chemical and atmospheric conditions bi-hourly over three different 24-h periods. Community composition of zooplankton export varied over the course of the day, and exports were most closely associated with wind directionality. Future studies of temporal variation in CCZE should incorporate wind conditions, especially in shallow systems where holomixis occurs frequently. Polymictic reservoirs are becoming increasingly common as the global pace of small dam construction quickens, making both the identification of factors influencing CCZE and the impact of zooplankton exports on local biodiversity and ecosystem function increasingly important to understand

Predicting Seasonal and Spatial Onset of cHABs in Polymictic Reservoirs

Cyanobacterial Harmful Algal Blooms (cHABS) are a naturally occurring but increasingly common phenomenon due to anthropogenic activities and climate change. cHABs reduce water quality by forming unsightly surface scums and sometimes producing algal matts on the surface of water bodies, reduce water quality, and in high densities can produce cyanotoxins that can harm humans, pets, and wildlife. Ecological forecasting of cHABs has proved elusive in part because the in-situ fluorometric methods currently employed for detecting cyanobacteria cells are subject to varied interference as water quality and the biotic community changes. In this study we seek to develop an ecological forecasting capability that overcomes both temporally and spatially derived in-situ fluorometric interferences. We obtained water samples at 26 polymictic reservoirs over a two-day period and at five polymictic reservoirs weekly during the summer of 2019. Collected water samples are being used for quantitative analysis of cyanobacterial cell densities by means of qPCR. We plan a data reduction technique (e.g. PCA, VIF screening, elastic-net regression as appropriate) followed by multivariate predictive model (e.g. multiple regression, ordination, discriminant analysis as appropriate)

Correlation Matrices of Cyanobacterial Bloom Predictors Varies Between Lakes

Under anthropogenically-altered conditions, cyanobacteria may form harmful algal blooms (cHABs) that can be toxic and disrupt ecosystem function. Developing tools to predict cHABs is an increasingly important task, but such tools have been difficult to develop. In our study, we contribute to the development of a predictive model for cHAB formation in the waters of southern New Jersey by statistically screening water quality data from five polymictic reservoirs that were sampled weekly from June through September 2019. The correlation structure of water quality variables differed between the reservoirs in a way that suggests that the mean correlation coefficient is elevated for reservoirs experiencing a cHAB. Some water quality variables are unlikely to be useful for predictive modeling, but among those that do have utility, those measurements were obtained under natural field conditions, semi-controlled conditions in the field, and controlled conditions in the lab. The number of principal components (PC axes) required to describe variation in the water quality data differed between reservoirs in a way that suggests reservoirs experiencing cHABs have less complex covariance structures. Collectively, these results indicate that predictive modeling of cHAB formation should be possible

Environmental Predictors of Zooplankton Biodiversity Across a Series of Polymictic Reservoirs

Zooplankton are an important part of lentic food webs because they connect photoautotrophic plankton to higher trophic levels. Past studies in 2016 and 2017 determined that zooplankton biodiversity is different across a polymictic reservoir series in Salem County NJ. In the study presented here, we confirmed a similar zooplankton biodiversity pattern in 2018 and we determine which physicochemical variables were good predictors of zooplankton biodiversity. Our results indicate that there are many physicochemical variables that are reliable predictors of zooplankton biodiversity across this polymictic reservoir series. Additional predictor variables are being developed to help us understand which environmental variables are most influential on zooplankton biodiversity and the influence of human land-use

Generation of a Kupffer Cell-evading Adenovirus for Systemic and Liver-directed Gene Transfer

As much as 90% of an intravenously (i.v.) injected dose of adenovirus serotype 5 (Ad5) is absorbed and destroyed by liver Kupffer cells. Viruses that escape these cells can then transduce hepatocytes after binding factor X (FX). Given that interactions with FX and Kupffer cells are thought to occur on the Ad5 hexon protein, we replaced its exposed hypervariable regions (HVR) with those from Ad6. When tested in vivo in BALB/c mice and in hamsters, the Ad5/6 chimera mediated \u3e10 times higher transduction in the liver. This effect was not due to changes in FX binding. Rather, Ad5/6 appeared to escape Kupffer cell uptake as evidenced by producing no Kupffer cell death in vivo, not requiring predosing in vivo, and being phagocytosed less efficiently by macrophages in vitro compared to Ad5. When tested as a helper-dependent adenovirus (Ad) vector, Ad5/6 mediated higher luciferase and factor IX transgene expression than either helper-dependent adenoviral 5 (HD-Ad5) or HD-Ad6 vectors. These data suggest that the Ad5/6 hexon-chimera evades Kupffer cells and may have utility for systemic and liver-directed therapies

Generation of a Kupffer Cell-evading Adenovirus for Systemic and Liver-directed Gene Transfer

As much as 90% of an intravenously (i.v.) injected dose of adenovirus serotype 5 (Ad5) is absorbed and destroyed by liver Kupffer cells. Viruses that escape these cells can then transduce hepatocytes after binding factor X (FX). Given that interactions with FX and Kupffer cells are thought to occur on the Ad5 hexon protein, we replaced its exposed hypervariable regions (HVR) with those from Ad6. When tested in vivo in BALB/c mice and in hamsters, the Ad5/6 chimera mediated \u3e10 times higher transduction in the liver. This effect was not due to changes in FX binding. Rather, Ad5/6 appeared to escape Kupffer cell uptake as evidenced by producing no Kupffer cell death in vivo, not requiring predosing in vivo, and being phagocytosed less efficiently by macrophages in vitro compared to Ad5. When tested as a helper-dependent adenovirus (Ad) vec- tor, Ad5/6 mediated higher luciferase and factor IX trans- gene expression than either helper-dependent adenoviral 5 (HD-Ad5) or HD-Ad6 vectors. These data suggest that the Ad5/6 hexon-chimera evades Kupffer cells and may have utility for systemic and liver-directed therapies

Expression capable library for studies of Neisseria gonorrhoeae, version 1.0

Background The sexually transmitted disease, gonorrhea, is a serious health problem in developed as well as in developing countries, for which treatment continues to be a challenge. The recent completion of the genome sequence of the causative agent, Neisseria gonorrhoeae, opens up an entirely new set of approaches for studying this organism and the diseases it causes. Here, we describe the initial phases of the construction of an expression-capable clone set representing the protein-coding ORFs of the gonococcal genome using a recombination-based cloning system. Results The clone set thus far includes 1672 of the 2250 predicted ORFs of the N. gonorrhoeae genome, of which 1393 (83%) are sequence-validated. Included in this set are 48 of the 61 ORFs of the gonococcal genetic island of strain MS11, not present in the sequenced genome of strain FA1090. L-arabinose-inducible glutathione-S-transferase (GST)-fusions were constructed from random clones and each was shown to express a fusion protein of the predicted size following induction, demonstrating the use of the recombination cloning system. PCR amplicons of each ORF used in the cloning reactions were spotted onto glass slides to produce DNA microarrays representing 2035 genes of the gonococcal genome. Pilot experiments indicate that these arrays are suitable for the analysis of global gene expression in gonococci. Conclusion This archived set of Gateway® entry clones will facilitate high-throughput genomic and proteomic studies of gonococcal genes using a variety of expression and analysis systems. In addition, the DNA arrays produced will allow us to generate gene expression profiles of gonococci grown in a wide variety of conditions. Together, the resources produced in this work will facilitate experiments to dissect the molecular mechanisms of gonococcal pathogenesis on a global scale, and ultimately lead to the determination of the functions of unknown genes in the genome

Evaluation of commercially available RNA amplification kits for RNA sequencing using very low input amounts of total RNA

This article includes supplemental data. Please visit http://www.fasebj.org to obtain this information.Multiple recent publications on RNA sequencing (RNA-seq) have demonstrated the power of next-generation sequencing technologies in whole-transcriptome analysis. Vendor-specific protocols used for RNA library construction often require at least 100 ng total RNA. However, under certain conditions, much less RNA is available for library construction. In these cases, effective transcriptome profiling requires amplification of subnanogram amounts of RNA. Several commercial RNA amplification kits are available for amplification prior to library construction for next-generation sequencing, but these kits have not been comprehensively field evaluated for accuracy and performance of RNA-seq for picogram amounts of RNA. To address this, 4 types of amplification kits were tested with 3 different concentrations, from 5 ng to 50 pg, of a commercially available RNA. Kits were tested at multiple sites to assess reproducibility and ease of use. The human total reference RNA used was spiked with a control pool of RNA molecules in order to further evaluate quantitative recovery of input material. Additional control data sets were generated from libraries constructed following polyA selection or ribosomal depletion using established kits and protocols. cDNA was collected from the different sites, and libraries were synthesized at a single site using established protocols. Sequencing runs were carried out on the Illumina platform. Numerous metrics were compared among the kits and dilutions used. Overall, no single kit appeared to meet all the challenges of small input material. However, it is encouraging that excellent data can be recovered with even the 50 pg input total RNA