Negative and Non-Positive Epidemiological Studies

Aim. To identify and discuss validity aspects on so called negative and non-positive studies. Methods. Arguments and examples are drawn from experiences in occupational health epidemiology regarding the interpretation of more or less equivocal study results. Results and conclusions. A negative study may be defined as showing a result that goes against the investigated hypothesis of an increased (or prevented) risk. Traditionally, studies with a risk estimate (relative risk or odds ratio) above but close to unity are also referred to as negative, given a narrow confidence interval (CI) that includes unity. A risk estimate above unity with the CI including unity is non-positive, however, but an estimate below unity with upper CI bond exceeding unity might be seen as possibly negative or non-negative. A weaker “significance” than usually required should perhaps be accepted when evaluating serious hazards. In contrast to positive studies, the negative and non-positive studies tend to escape criticism in spite of questionable validity that may have obscured existing risks (or preventive effects). Even stronger arguments can be made in criticising negative and non-positive studies than positive studies, for example, regarding selection phenomena, and observational problems regarding exposure and outcome. Negative confounding should be considered although usually weak. In case-control studies, so-called over-matching may obscure an existing risk as could the “healthy worker effect” in cohort studies. Small scale non-positive studies should be made available for meta-analyses and when considering studies that do not convincingly show a risk; those who are exposed should be given the “benefit of the doubt.”     

Pygidialporen bei marinen Enchytraeidae (Oligochaeta)

Coelo-rectal pygidial pores were found in the anal area of marine enchytraeids (Lumbricillus lineatus, Marionina spicula, Enchytraeus albidus) from the mud-flats of the Isle of Sylt. The structure and function of these pores have been discussed.     

Automated counting of mammalian cell colonies by means of a flat bed scanner and image processing.

BACKGROUND: Clonogenic assays are used frequently to measure the cell killing and mutagenic effects of radiation and other agents. Clonogenic assays carried out manually are tedious and time-consuming and involve a significant element of subjectivity. However, several commercial automatic colony counters are available. Based on CCD video imaging and image analysis they are relatively expensive and can analyze only one petri dish at a time. METHOD: We have developed a cheaper and more efficient device, which employs a flat bed scanner to image 12 60-mm petri dishes at a time. Two major problems in automated colony counting are the clustering of colonies and edge effects. By using standard image analysis and implementing an inflection point algorithm, these problems were greatly diminished. The resulting system was compared with two manual colony counts, as well as with automated counts with the Oxford Optronix ColCount colony counter for cell lines V79 and HaCaT. RESULTS: Comparisons assuming the manual counts to be correct showed that our automatic counter was slightly more accurate than the commercial unit. CONCLUSIONS: As a whole, our automated colony counter performed significantly better than the commercial unit with regard to processing time, cost and accuracy.     

Sequence comparison and mutational analysis of elements that may be involved in the regulation of DNA synthesis in HIV-1

The large number of sequenced clones of HIV-1 and related viruses made it possible to indicate conserved elements with potential regulatory or structural functions. Such analysis was combined with directed mutagenesis in order to investigate the importance of elements that may influence the initiation of plus-strand DNA synthesis. The main site for plus-strand initiation is a polypurine tract near the 3′ end of the viral RNA (the 3′ PPT). An exact copy of this PPT is located in the middle of the genome (the internal PPT). Upstream from the internal PPT there is an inverted repeat. Mutants designed to upset the internal PPT (i.e., purine to pyrimidine changes), as well as mutants designed to abolish the potential stem-loop formation (changes around the internal PPT or in the upstream inverted repeat) both resulted in viruses with a reduced ability to replicate. Upsetting the stem-loop formation was, however, less harmful than changing the polypurine nature of the PPT. Changing a conserved T on the 3′ side of the PPT to a C did not affect the phenotype. Presented at the NATO Advanced Research Workshop onGenome Organization and Evolution, Spetsai, Greece, 16–22 September 1992     

Characterization of the inhibitory effect of glucocorticoids on the DNA replication of adult rat hepatocytes growing at various cell densities

Dexamethasone inhibited the basal and EGF-stimulated DNA synthesis of adult rat hepatocytes in primary culture. The inhibition was glucocorticoid-specific: It was shown by dexamethasone and hydrocortisone, but not by progesterone, testosterone, or estradiol; and was counteracted by the glucocorticoid antagonist RU-38486 in a concentration-dependent manner. Dexamethasone acted by decreasing the rate of entry into S-phase (kG1/S), while cell cycle parameters were unaffected. The steroid was able to decrease the kG1/S severalfold even when added more than 20 hr after EGF, half-maximal effect occurring 11 hr after the addition of dexamethasone. Densely populated areas were much more sensitive to the inhibition by dexamethasone than sparsely populated areas within the same culture dish: A moderate (10 nM) concentration of dexamethasone nearly abolished the DNA synthesis in densely populated areas of hepatocyte cultures with only marginal effect on sparsely populated cells.     

Effects of acorn abundance on density dependence in a Japanese wood mouse (Apodemus speciosus) population

We analysed the effects of Quercus crispula acorn abundance on the density dependence of the large Japanese wood mouse Apodemus speciosus using time series data (1992–2007). The data were obtained in a forest in northern Hokkaido, Japan, by live-trapping rodents and directly counting acorns on the ground. Acorn abundance in one year clearly influenced the abundance of wood mice in the following year in all models examined based on the Gompertz and Ricker model; in addition, the abundance of wood mice had effects on the population. Acorn abundance influenced the strength of density dependence (intraspecific competition) of the wood mouse population. When the abundance of acorns was high, density dependence was relaxed, and as a result the equilibrium density at which the population growth rate decreased to zero became higher. Those effects of acorn abundance were regarded as a nonlinear perturbation effect (sensu Royama 1992). The nonlinearity of density dependence was also detected; higher densities had stronger effects on population growth rates. Electronic supplementary material The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Microbial Community in Black Rust Exposed to Hot Ridge Flank Crustal Fluids

During Integrated Ocean Drilling Program Expedition 301, we obtained a sample of black rust from a circulation obviation retrofit kit (CORK) observatory at a borehole on the eastern flank of Juan de Fuca Ridge. Due to overpressure, the CORK had failed to seal the borehole. Hot fluids from oceanic crust had discharged to the overlying bottom seawater and resulted in the formation of black rust analogous to a hydrothermal chimney deposit. Both culture-dependent and culture-independent analyses indicated that the black-rust-associated community differed from communities reported from other microbial habitats, including hydrothermal vents at seafloor spreading centers, while it shared phylotypes with communities previously detected in crustal fluids from the same borehole. The most frequently retrieved sequences of bacterial and archaeal 16S rRNA genes were related to the genera Ammonifex and Methanothermococcus, respectively. Most phylotypes, including phylotypes previously detected in crustal fluids, were isolated in pure culture, and their metabolic traits were determined. Quantification of the dissimilatory sulfite reductase (dsrAB) genes, together with stable sulfur isotopic and electron microscopic analyses, strongly suggested the prevalence of sulfate reduction, potentially by the Ammonifex group of bacteria. Stable carbon isotopic analyses suggested that the bulk of the microbial community was trophically reliant upon photosynthesis-derived organic matter. This report provides important insights into the phylogenetic, physiological, and trophic characteristics of subseafloor microbial ecosystems in warm ridge flank crusts.     

GH–IGF system regulation of attenuated muscle growth and lipolysis in Atlantic salmon reared at elevated sea temperatures

Growth regulation in adult Atlantic salmon (1.6 kg) was investigated during 45 days in seawater at 13, 15, 17, and 19 °C. We focused on feed intake, nutrient uptake, nutrient utilization, and endocrine regulation through growth hormone (GH), insulin-like growth factors (IGF), and IGF-binding proteins (IGFBP). During prolonged thermal exposure, salmon reduced feed intake and growth. Feed utilization was reduced at 19 °C after 45 days compared with fish at lower temperatures, and body lipid storage was depleted with increasing water temperature. Although plasma IGF-1 concentrations did not change, 32-Da and 43-kDa IGFBP increased in fish reared at ≤17 °C, and dropped in fish reared at 19 °C. Muscle igf1 mRNA levels were reduced at 15 and 45 days in fish reared at 15, 17, and 19 °C. Muscle igf2 mRNA levels did not change after 15 days in response to increasing temperature, but were reduced after 45 days. Although liver igf2 mRNA levels were reduced with increasing temperatures after 15 and 45 days, temperature had no effect on igf1 mRNA levels. The liver igfbp2b mRNA level, which corresponds to circulating 43-kDa IGFBP, exhibited similar responses after 45 days. IGFBP of 23 kDa was only detected in plasma in fish reared at 17 °C, and up-regulation of the corresponding igfbp1b gene indicated a time-dependent catabolic response, which was not observed in fish reared at 19 °C. However, higher muscle ghr mRNA levels were detected in fish at 17 and 19 °C than in fish at lower temperatures, indicating lipolytic regulation in muscle. These results show that the reduction of muscle growth in large salmon is mediated by decreased igf1 and igf2 mRNA levels in addition to GH-associated lipolytic action to cope with prolonged thermal exposure. Accordingly, 13 °C appears to be a more optimal temperature for the growth of adult Atlantic salmon at sea.     

Environmental enrichment promotes neural plasticity and cognitive ability in fish

Different kinds of experience during early life can play a significant role in the development of an animal''s behavioural phenotype. In natural contexts, this influences behaviours from anti-predator responses to navigation abilities. By contrast, for animals reared in captive environments, the homogeneous nature of their experience tends to reduce behavioural flexibility. Studies with cage-reared rodents indicate that captivity often compromises neural development and neural plasticity. Such neural and behavioural deficits can be problematic if captive-bred animals are being reared with the intention of releasing them as part of a conservation strategy. Over the last decade, there has been growing interest in the use of environmental enrichment to promote behavioural flexibility in animals that are bred for release. Here, we describe the positive effects of environmental enrichment on neural plasticity and cognition in juvenile Atlantic salmon (Salmo salar). Exposing fish to enriched conditions upregulated the forebrain expression of NeuroD1 mRNA and improved learning ability assessed in a spatial task. The addition of enrichment to the captive environment thus promotes neural and behavioural changes that are likely to promote behavioural flexibility and improve post-release survival.