Studies on the effects of subzero temperatures on the viability of spores of Aspergillus flavus. I. The effect of rate of warming

1. The survival of spores of Aspergillus flavus suspended in distilled water and cooled rapidly to –70 to –75°C. was found to depend primarily on the rate of subsequent warming of the frozen suspension. Only 7 per cent of the spores germinated following slow warming at 0.9°C. per minute, whereas about 75 per cent germinated following rapid warming at 700°C. per minute. 2. Viability was dependent on the rate at which the suspensions warmed from –70 to 0°C. (subzero warming), but was not dependent on the rate of thawing of the frozen water in which the spores were suspended. 3. The logarithm of the percentage of germination appeared to be a linear function of the logarithm of the rate of subzero warming when spores were warmed at rates ranging from 0.12 to 1000°C. per minute. 4. The lethal effects of slow warming from –70 to 0°C. were more pronounced between about –20 and 0°C. than between –70 and –20°C. In the former range of temperatures, the percentage of germination decreased sharply as slow warming progressed towards 0°C. 5. Slow warming from –70 to 0°C. was more harmful to the spores than was a 1 or 2 hour exposure to constant temperatures between –70 and 0°C. 6. Slow warming was found to be more harmful than rapid warming when spores were suspended in horse serum, 0.16 molal sodium chloride, or 0.29 molal sucrose as well as in distilled water.     

Cold-Induced Vasodilation during Single Digit Immersion in 0°C and 8°C Water in Men and Women

The present study compared the thermal responses of the finger to 0 and 8°C water immersion, two commonly used temperatures for cold-induced vasodilation (CIVD) research. On two separate and counterbalanced occasions 15 male and 15 female participants immersed their index finger in 20°C water for 5 min followed by either 0 or 8°C water for 30 min. Skin temperature, cardiovascular and perceptual data were recorded. Secondary analyses were performed between sexes and comparing 0.5, 1 and 4°C CIVD amplitude thresholds. With a 0.5°C threshold, CIVD waves were more prevalent in 8°C (2 (1 – 3) than in 0°C (1.5 (0 – 3)), but the amplitude was lower (4.0 ± 2.3 v 9.2 ± 4.0°C). Mean, minimum and maximum finger temperatures were lower in 0°C during the 30 min immersion, and CIVD onset and peak time occurred later in 0°C. Thermal sensation was lower and pain sensation was higher in 0°C. There were no differences between males and females in any of the physiological or CIVD data with the exception of SBP, which was higher in males. Females reported feeling higher thermal sensations in 8°C and lower pain sensations in 0°C and 8°C compared to males. Fewer CIVD responses were observed when using a 4°C (1 (0 – 3)) threshold to quantify a CIVD wave compared to using a 1°C (2 (0 – 3)) or 0.5°C (2 (0 – 3)) amplitude. In conclusion, both 0 and 8 °C can elicit CIVD but 8°C may be more suitable when looking to optimise the number of CIVD waves while minimising participant discomfort. The CIVD response to water immersion does not appear to be influenced by sex. Researchers should consider the amplitude threshold that was used to determine a CIVD wave when interpreting previous data.     

CONCERNING THE HEREDITARY ADAPTATION OF ORGANISMS TO HIGHER TEMPERATURE

1. Imagos of Drosophila raised at temperatures of from 12–28.5°C. when placed at any temperature from 15–32.5°C. produce eggs which develop normally at these temperatures. 2. Imagos raised at temperatures of from 29–32.5° and then kept permanently within these temperatures produce eggs which do not develop. 3. Imagos raised at from 28.5–32.5°C. and then placed at temperatures of from 12–25°C. produce eggs which develop normally. 4. Imagos raised at from 28.5–32.5°C. placed at 15–25°C. for 24 hours or longer and then put back into a temperature of from 28.5–32.5°C., produce eggs which will develop at the latter temperature. 5. There is no evidence of any hereditary adaptation to higher temperatures.     

Cold-sensitive mutants of Taq DNA polymerase provide a hot start for PCR

Although the thermophilic bacterium Thermus aquaticus grows optimally at 70°C and cannot grow at moderate temperatures, its DNA polymerase I has significant activity at 20–37°C. This activity is a bane to some PCRs, since it catalyzes non-specific priming. We report mutations of Klentaq (an N-terminal deletion variant) DNA polymerase that have markedly reduced activity at 37°C yet retain apparently normal activity at 68°C and resistance at 95°C. The first four of these mutations are clustered on the outside surface of the enzyme, nowhere near the active site, but at the hinge point of a domain that has been proposed to move at each cycle of nucleotide incorporation. We show that the novel cold-sensitive mutants can provide a hot start for PCR and exhibit slightly improved fidelity.     

Patterns of odour emission,thermogenesis and pollinator activity in cones of an African cycad: what mechanisms apply?

Background and Aims

Ontogenetic patterns of odour emissions and heating associated with plant reproductive structures may have profound effects on insect behaviour, and consequently on pollination. In some cycads, notably Macrozamia, temporal changes in emission of specific odour compounds and temperature have been interpreted as a ‘push–pull’ interaction in which pollinators are either attracted or repelled according to the concentration of the emitted volatiles. To establish which mechanisms occur in the large Encephalartos cycad clade, the temporal patterns of volatile emissions, heating and pollinator activity of cones of Encephalartos villosus in the Eastern Cape (EC) and KwaZulu Natal (KZN) of South Africa were investigated.

Methods and Key Results

Gas chromatography–mass spectrometry (GC-MS) analyses of Encephalartos villosus cone volatiles showed that emissions, dominated by eucalyptol and 2-isopropyl-3-methoxypyrazine in EC populations and (3E)-1,3-octadiene and (3E,5Z)-1,3,5-octatriene in the KZN populations, varied across developmental stages but did not vary significantly on a daily cycle. Heating in male cones was higher at dehiscence than during pre- and post-dehiscence, and reached a maximum at about 1830 h when temperatures were between 7·0 and 12·0 °C above ambient. Daily heating of female cones was less pronounced and reached a maximum at about 1345 h when it was on average between 0·9 and 3·0 °C above ambient. Insect abundance on male cones was higher at dehiscence than at the other stages and significantly higher in the afternoon than in the morning and evening.

Conclusions

There are pronounced developmental changes in volatile emissions and heating in E. villosus cones, as well as strong daily changes in thermogenesis. Daily patterns of volatile emissions and pollinator abundance in E. villosus are different from those observed in some Macrozamia cycads and not consistent with the push–pull pattern as periods of peak odour emission do not coincide with mass exodus of insects from male cones.     

Plant Morphological and Biochemical Responses to Field Water Deficits: III. Effect of Foliage Temperature on the Potential Activity of Glutathione Reductase

Activity of glutathione reductase has been related to stress tolerance; however, these enzyme assays are generally conducted at 25°C. Foliage temperature varies greatly in the field in response to soil water availability and ambient conditions and this may affect enzyme response. This study was conducted to determine the effect of changing foliage temperature on glutathione reductase activity of wheat under field conditions. Wheat leaf glutathione reductase was purified and the temperature response of the enzyme was determined at 2.5°C intervals between 12.5 and 45°C. These data, in conjunction with continuous measurements of field-grown wheat foliage temperatures, were used to compare the temperature-related changes in potential glutathione reductase activities in water stressed and control plants. Assuming saturating substrate levels, the results indicate that early in the season the daily potential enzyme activity of the irrigated and stressed plants could never have reached the daily activity predicted from the 25°C (room temperature) measurements. Later in the season, the daily potential activity of the irrigated plants was lower, and the daily potential activity of the stressed plants was higher, than the activities predicted from the 25°C determinations. These results suggest that a better understanding of the regulation of plant metabolism will be obtained by linking continuous temperature measurements of plant foliage with enzyme responses to temperature.     

Recombinant production and biochemical characterization of a hyperthermostable α-glucan/maltodextrin phosphorylase from Pyrococcusfuriosus

Alpha-glucan phosphorylase catalyzes the reversible cleavage of α-1-4-linked glucose polymers into α-D-glucose-1-phosphate. We report the recombinant production of an α-glucan/maltodextrin phosphorylase (PF1535) from a hyperthermophilic archaeon, Pyrococcus furiosus, and the first detailed biochemical characterization of this enzyme from any archaeal source using a mass-spectrometry-based assay. The apparent 98 kDa recombinant enzyme was active over a broad range of temperatures and pH, with optimal activity at 80 °C and pH 6.5–7. This archaeal protein retained its complete activity after 24 h at 80 °C in Tris-HCl buffer. Unlike other previously reported phosphorylases, the Ni-affinity column purified enzyme showed broad substrate specificity in both the synthesis and degradation of maltooligosaccharides. In the synthetic direction of the enzymatic reaction, the lowest oligosaccharide required for the chain elongation was maltose. In the degradative direction, the archaeal enzyme can produce glucose-1-phosphate from maltotriose or longer maltooligosaccharides including both glycogen and starch. The specific activity of the enzyme at 80 °C in the presence of 10 mM maltoheptaose and at 10 mg ml^–1 glycogen concentration was 52 U mg^–1 and 31 U mg^–1, respectively. The apparent Michaelis constant and maximum velocity for inorganic phosphate were 31 ± 2 mM and 0.60 ± 0.02 mM min^–1 µg^–1, respectively. An initial velocity study of the enzymatic reaction indicated a sequential bi-bi catalytic mechanism. Unlike the more widely studied mammalian glycogen phosphorylase, the Pyrococcus enzyme is active in the absence of added AMP.     

A Novel Method Applied in Determination and Assessment of Trace Amount of Lead and Cadmium in Rice from Four Provinces,China

Heavy metal contamination of soils or water can lead to excessive lead (Pb) and cadmium (Cd) levels in rice. As cumulative poisons, consumption of Pb and Cd in contaminated rice may cause many toxic effects in humans. In the present study, Pb and Cd levels in rice samples from Hubei, Jiangxi, Heilongjiang, and Guangdong provinces in China were analyzed by cloud point extraction and graphite furnace atomic absorption spectrometry (GFAAS). The heavy metals in the rice samples were reacted with 8-quinolinol to form a complex at pH 9.0 and 40°C. Analytes were quantitatively extracted to a surfactant-rich phase (Triton X-45) after centrifugation and analyzed by GFAAS. The effects of experimental conditions, including pH, concentration of reagents, and equilibration time and temperature, on cloud point extraction were optimized efficiently using Plackett–Burman and Box–Behnken experimental designs. Under the optimum conditions, good linearity was observed in the concentration ranges of 0.5–5 µg/L for Pb and 0.05–0.50 µg/L for Cd. The limits of detection were 0.043 µg/L for Pb with a concentration factor of 24.2 in a 10 mL sample and 0.018 µg/L for Cd with a concentration factor of 18.4 in a 10 mL sample. Twenty rice samples from four provinces were analyzed successfully, and the mean levels of Pb and Cd in the rice were all below their maximum allowable concentrations in China. Comparing the tolerable daily intakes given by FAO/WHO with the mean estimated daily intakes; Pb and Cd mean daily intake through rice consumption were 0.84 µg/kg bw/day and 0.40 µg/kg bw/day, which were lower than the tolerable daily intakes.     

Biofilm Production Potential of Salmonella Serovars Isolated from Chickens in North West Province,South Africa

Bacterial biofilms have recently gained considerable interest in the food production and medical industries due to their ability to resist destruction by disinfectants and other antimicrobials. Biofilms are extracellular polymer matrices that may enhance the survival of pathogens even when exposed to environmental stress. The effect of incubation temperatures (25°C, 37°C, and 40°C) and Salmonella serotype on biofilm-forming potentials was evaluated. Previously typed Salmonella serotypes (55) isolated from the gut of chickens were accessed for biofilms formation using a standard assay. Salmonella Typhimurium ATCC 14028^TM and Salmonella Enteritidis ATCC 13076^TM (positive controls), Escherichia coli (internal control) and un-inoculated Luria Bertani (LB) broth (negative control) were used. The isolates formed no biofilm (11.86–13.56%), weak (11.86–45.76%), moderate (18.64–20.34%), strong biofilms (23.73–54.24%) across the various temperatures investigated. Serotypes, Salmonella Heidelberg and Salmonella Weltevreden were the strongest biofilm formers at temperatures (25°C, 37°C, and 40°C, respectively). The potential of a large proportion (80%) of Salmonella serotypes to form biofilms increased with increasing incubation temperatures but decreased at 40°C. Findings indicate that average temperature favours biofilm formation by Salmonella serotypes. However, the influence of incubation temperature on biofilm formation was greater when compared to serotype. A positive correlation exists between Salmonella biofilm formed at 25°C, 37°C and 40°C (p ≥ 0.01). The ability of Salmonella species to form biofilms at 25°C and 37°C suggests that these serotypes may present severe challenges to food-processing and hospital facilities.Key words: Salmonella, biofilm, biofilm production potential, crystal violet microtitre     

SUCROSE INVERSION BY BAKERS' YEAST AS A FUNCTION OF TEMPERATURE

Inversion of sucrose by bakers'' yeast follows the same course as inversion catalyzed by yeast invertase. Rate of inversion increases exponentially with temperature; the temperature characteristic in the Arrhenius equation is 10,700 below 13–17°C., and 8,300 above that temperature. Temperature inactivation occurs above 40°C. The effects of temperature upon rate of inversion were the same using Fleischmann''s yeast cake, the same yeast killed with toluene, and a pure strain (G. M. No. 21062) of bakers'' yeast. The last differed from the other two only in the fact that its critical temperature was 13°C. as compared with 17°C. for the others. The catalytic inversion is associated with enzyme activity inside the cell, not in the medium, and is independent of any vital processes inside the cell such as respiration and fermentation. Since invertase activity is the same inside the cell as it is after extraction, it appears possible to relate the temperature characteristics for physiological processes to the catalytic chemical systems which determine their rate. At least two enzymes are capable of inverting sucrose in the yeast cell. The familiar yeast invertase (µ = 10,700) is active below 13–17°C. while a second enzyme (M = 8,300) plays the dominant role above that temperature.     

Design,synthesis, anticancer evaluation,and molecular modelling studies of novel tolmetin derivatives as potential VEGFR-2 inhibitors and apoptosis inducers

Novel tolmetin derivatives 5a–f to 8a–c were designed, synthesised, and evaluated for antiproliferative activity by NCI (USA) against a panel of 60 tumour cell lines. The cytotoxic activity of the most active tolmetin derivatives 5b and 5c was examined against HL-60, HCT-15, and UO-31 tumour cell lines. Compound 5b was found to be the most potent derivative against HL-60, HCT-15, and UO-31 cell lines with IC₅₀ values of 10.32 ± 0.55, 6.62 ± 0.35, and 7.69 ± 0.41 µM, respectively. Molecular modelling studies of derivative 5b towards the VEGFR-2 active site were performed. Compound 5b displayed high inhibitory activity against VEGFR-2 (IC₅₀ = 0.20 µM). It extremely reduced the HUVECs migration potential exhibiting deeply reduced wound healing patterns after 72 h. It induced apoptosis in HCT-15 cells (52.72-fold). This evidence was supported by an increase in the level of apoptotic caspases-3, -8, and -9 by 7.808-, 1.867-, and 7.622-fold, respectively. Compound 5b arrested the cell cycle in the G0/G1 phase. Furthermore, the ADME studies showed that compound 5b possessed promising pharmacokinetic properties.     

Purification and Properties of Two Thermostable Alkaline Xylanases from an Alkaliphilic Bacillus sp.

Two xylanases, designated XylA and XylB, were purified from the culture supernatant of the alkaliphilic Bacillus sp. strain AR-009. The molecular masses of the two enzymes were estimated to be 23 kDa (XylA) and 48 kDa (XylB) by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. The optimum pHs for activity were 9 for XylA and 9 to 10 for XylB. The temperature optima for the activity of XylA were 60°C at pH 9 and 70°C at pH 8. XylB was optimally active at 75°C at pH 9 and 70°C at pH 8. Both enzymes were stable in a broad pH range and showed good stability when incubated at 60 and 65°C in pH 8 and 9 buffers.     

BOUND WATER IN MUSCLE

1. The amount of free unfrozen water, i.e. water acting as normal solvent, in frog''s muscle at temperatures below the initial freezing-point has been calculated from the vapour pressure isotherm of the muscle. 2. Significant amounts of free water are present at –20°C. The total amount of unfrozen water at –20°C. cannot, therefore, be taken as a measure of the bound water in muscle. 3. The calculated values of free water, when compared with experimentally determined values of total unfrozen water, indicate that the amount of bound water in muscle at various temperatures is small. 4. A temperature considerably below –20°C., roughly between –40° and –60°C., is required to freeze completely the free water in muscle.     

Interference competition between wolves and coyotes during variable prey abundance

Interference competition occurs when two species have similar resource requirements and one species is dominant and can suppress or exclude the subordinate species. Wolves (Canis lupus) and coyotes (C. latrans) are sympatric across much of their range in North America where white‐tailed deer (Odocoileus virginianus) can be an important prey species. We assessed the extent of niche overlap between wolves and coyotes using activity, diet, and space use as evidence for interference competition during three periods related to the availability of white‐tailed deer fawns in the Upper Great Lakes region of the USA. We assessed activity overlap (Δ) with data from accelerometers onboard global positioning system (GPS) collars worn by wolves (n = 11) and coyotes (n = 13). We analyzed wolf and coyote scat to estimate dietary breadth (B) and food niche overlap (α). We used resource utilization functions (RUFs) with canid GPS location data, white‐tailed deer RUFs, ruffed grouse (Bonasa umbellus) and snowshoe hare (Lepus americanus) densities, and landscape covariates to compare population‐level space use. Wolves and coyotes exhibited considerable overlap in activity (Δ = 0.86–0.92), diet (B = 3.1–4.9; α = 0.76–1.0), and space use of active and inactive RUFs across time periods. Coyotes relied less on deer as prey compared to wolves and consumed greater amounts of smaller prey items. Coyotes exhibited greater population‐level variation in space use compared to wolves. Additionally, while active and inactive, coyotes exhibited greater selection of some land covers as compared to wolves. Our findings lend support for interference competition between wolves and coyotes with significant overlap across resource attributes examined. The mechanisms through which wolves and coyotes coexist appear to be driven largely by how coyotes, a generalist species, exploit narrow differences in resource availability and display greater population‐level plasticity in resource use.     

Non-Instrumented Incubation of a Recombinase Polymerase Amplification Assay for the Rapid and Sensitive Detection of Proviral HIV-1 DNA

Sensitive diagnostic tests for infectious diseases often employ nucleic acid amplification technologies (NAATs). However, most NAAT assays, including many isothermal amplification methods, require power-dependent instrumentation for incubation. For use in low resource settings (LRS), diagnostics that do not require consistent electricity supply would be ideal. Recombinase polymerase amplification (RPA) is an isothermal amplification technology that has been shown to typically work at temperatures ranging from 25–43°C, and does not require a stringent incubation temperature for optimal performance. Here we evaluate the ability to incubate an HIV-1 RPA assay, intended for use as an infant HIV diagnostic in LRS, at ambient temperatures or with a simple non-instrumented heat source. To determine the range of expected ambient temperatures in settings where an HIV-1 infant diagnostic would be of most use, a dataset of the seasonal range of daily temperatures in sub Saharan Africa was analyzed and revealed ambient temperatures as low as 10°C and rarely above 43°C. All 24 of 24 (100%) HIV-1 RPA reactions amplified when incubated for 20 minutes between 31°C and 43°C. The amplification from the HIV-1 RPA assay under investigation at temperatures was less consistent below 30°C. Thus, we developed a chemical heater to incubate HIV-1 RPA assays when ambient temperatures are between 10°C and 30°C. All 12/12 (100%) reactions amplified with chemical heat incubation from ambient temperatures of 15°C, 20°C, 25°C and 30°C. We also observed that incubation at 30 minutes improved assay performance at lower temperatures where detection was sporadic using 20 minutes incubation. We have demonstrated that incubation of the RPA HIV-1 assay via ambient temperatures or using chemical heaters yields similar results to using electrically powered devices. We propose that this RPA HIV-1 assay may not need dedicated equipment to be a highly sensitive tool to diagnose infant HIV-1 in LRS.     

Changes in the Enzymes for Fatty Acid Synthesis and Desaturation during Acclimation of Developing Soybean Seeds to Altered Growth Temperature

Temperature-induced changes in the enzymes for fatty acid synthesis and desaturation were studied in developing soybean seeds (Glycine max L. var Williams 82). Changes were induced by culture of the seed pods for 20 hours in liquid media at 20, 25, or 35°C. Linoleoyl and oleoyl desaturases were 94 and 10 times as active, respectively, in seeds cultured at 20°C as those cultured at 25°C. Both desaturases had negligible activity in seeds cultured at 35°C compared to seeds cultured at 20°C. Though less dramatic, other enzymes also showed differences in activity after 20 hours in culture at 20, 25, or 35°C. Stearoyl-acyl carrier protein (ACP) desaturase and CDP-choline:diacylglycerol phosphorylcholine transferase were most active in preparations from 20°C cultures. Activities were twofold lower at 25°C and a further threefold lower in 35°C cultures. Cultures from 25 and 35°C had 60 and 40%, respectively, of the phosphorylcholine:CTP cytidylyl transferase activity present in cultures grown at 20°C. Fatty acid synthetase, malonyl-coenzyme A:ACP transacylase, palmitoyl-ACP elongation, and choline kinase were not significantly altered by culture temperature. These data suggest that the enzymes for fatty acid desaturation and phosphatidylcholine synthesis can be rapidly modulated in response to altered growth temperatures, while the enzymes for fatty acid synthesis and elongation are not.     

Impacts of growth form and phylogenetic relatedness on seed germination: A large‐scale analysis of a subtropical regional flora

Plant regeneration strategy plays a critical role in species survival and can be used as a proxy for the evolutionary response of species to climate change. However, information on the effects of key plant traits and phylogenetic relatedness on seed germination is limited at large regional scales that vary in climate. To test the hypotheses that phylogenetic niche conservatism plays a critical force in shaping seed ecophysiological traits across species, and also drives their response to climatic fluctuation, we conducted a controlled experiment on seed germination and determined the percentage and rate of germination for 249 species in subtropical China under two temperature regimes (i.e., daily 25°C; daily alternating 25/15°C for each 12 hr). Germination was low with a skewed distribution (mean = 38.9% at 25°C, and 43.3% at 25/15°C). One fifth of the species had low (<10%) and slow (4–30 days) germination, and only a few (8%) species had a high (>80%) and rapid (1.2–6.6 days) germination. All studied plant traits (including germination responses) showed a significant phylogenetic signal, with an exception of seed germination percentage under the alternating temperature scenario. Generalized linear models (GLMs) and phylogenetic generalized estimation equations (GEEs) demonstrated that growth form and seed dispersal mode were strong drivers of germination. Our experimental study highlights that integrating plant key traits and phylogeny is critical to predicting seed germination response to future climate change.     

Determination of Femoral Neck Angle and Torsion Angle Utilizing a Novel Three-Dimensional Modeling and Analytical Technology Based on CT Datasets

Introduction

Exact knowledge of femoral neck inclination and torsion angles is important in recognizing, understanding and treating pathologic conditions in the hip joint. However, published results vary widely between different studies, which indicates that there are persistent difficulties in carrying out exact measurements.

Methods

A three dimensional modeling and analytical technology was used for the analysis of 1070 CT datasets of skeletally mature femurs. Individual femoral neck angles and torsion angles were precisely computed, in order to establish whether gender, age, body mass index and ethnicity influence femoral neck angles and torsion angles.

Results

The median femoral neck angle was 122.2° (range 100.1–146.2°, IQR 117.9–125.6°). There are significant gender (female 123.0° vs. male 121.5°; p = 0.007) and ethnic (Asian 123.2° vs. Caucasian 121.9°; p = 0.0009) differences. The median femoral torsion angle was 14.2° (-23.6–48.7°, IQR 7.4–20.4°). There are significant gender differences (female 16.4° vs. male 12.1°; p = 0.0001). Femoral retroversion was found in 7.8% of the subjects.

Conclusion

Precise femoral neck and torsion angles were obtained in over one thousand cases. Systematic deviations in measurement due to human error were eliminated by using automated high accuracy morphometric analysis. Small but significant gender and ethnic differences were found in femoral neck and torsion angles.     

Multimodal Distribution of Human Cold Pain Thresholds

BackgroundIt is assumed that different pain phenotypes are based on varying molecular pathomechanisms. Distinct ion channels seem to be associated with the perception of cold pain, in particular TRPM8 and TRPA1 have been highlighted previously. The present study analyzed the distribution of cold pain thresholds with focus at describing the multimodality based on the hypothesis that it reflects a contribution of distinct ion channels.MethodsCold pain thresholds (CPT) were available from 329 healthy volunteers (aged 18 – 37 years; 159 men) enrolled in previous studies. The distribution of the pooled and log-transformed threshold data was described using a kernel density estimation (Pareto Density Estimation (PDE)) and subsequently, the log data was modeled as a mixture of Gaussian distributions using the expectation maximization (EM) algorithm to optimize the fit.ResultsCPTs were clearly multi-modally distributed. Fitting a Gaussian Mixture Model (GMM) to the log-transformed threshold data revealed that the best fit is obtained when applying a three-model distribution pattern. The modes of the identified three Gaussian distributions, retransformed from the log domain to the mean stimulation temperatures at which the subjects had indicated pain thresholds, were obtained at 23.7 °C, 13.2 °C and 1.5 °C for Gaussian #1, #2 and #3, respectively.ConclusionsThe localization of the first and second Gaussians was interpreted as reflecting the contribution of two different cold sensors. From the calculated localization of the modes of the first two Gaussians, the hypothesis of an involvement of TRPM8, sensing temperatures from 25 – 24 °C, and TRPA1, sensing cold from 17 °C can be derived. In that case, subjects belonging to either Gaussian would possess a dominance of the one or the other receptor at the skin area where the cold stimuli had been applied. The findings therefore support a suitability of complex analytical approaches to detect mechanistically determined patterns from pain phenotype data.     

Activity Patterns of Eurasian Lynx Are Modulated by Light Regime and Individual Traits over a Wide Latitudinal Range

The activity patterns of most terrestrial animals are regarded as being primarily influenced by light, although other factors, such as sexual cycle and climatic conditions, can modify the underlying patterns. However, most activity studies have been limited to a single study area, which in turn limit the variability of light conditions and other factors. Here we considered a range of variables that might potentially influence the activity of a large carnivore, the Eurasian lynx, in a network of studies conducted with identical methodology in different areas spanning latitudes from 49°7′N in central Europe to 70°00′N in northern Scandinavia. The variables considered both light conditions, ranging from a day with a complete day–night cycle to polar night and polar day, as well as individual traits of the animals. We analysed activity data of 38 individual free-ranging lynx equipped with GPS-collars with acceleration sensors, covering more than 11,000 lynx days. Mixed linear additive models revealed that the lynx activity level was not influenced by the daily daylight duration and the activity pattern was bimodal, even during polar night and polar day. The duration of the active phase of the activity cycle varied with the widening and narrowing of the photoperiod. Activity varied significantly with moonlight. Among adults, males were more active than females, and subadult lynx were more active than adults. In polar regions, the amplitude of the lynx daily activity pattern was low, likely as a result of the polycyclic activity pattern of their main prey, reindeer. At lower latitudes, the basic lynx activity pattern peaked during twilight, corresponding to the crepuscular activity pattern of the main prey, roe deer. Our results indicated that the basic activity of lynx is independent of light conditions, but is modified by both individual traits and the activity pattern of the locally most important prey.