Temperature-induced oviposition in the brachyuran crab Cancer setosus along a latitudinal cline: Aquaria experiments and analysis of field-data

Ovigerous females of Cancer setosus are present year-round throughout most of its wide range along the Peruvian/Chilean Pacific coast (2°S-46°S). However, their number of egg-masses produced per year remains speculative and as such has neither been considered in latitudinal comparisons of reproduction, nor for its fisheries management. In order to reveal the effect of temperature on egg-mass production and egg-development, female C. setosus were held in through-flow aquaria under natural seasonal temperature conditions (16-23 °C) in Antofagasta (23°S), Northern Chile (05/2005-03/2006; 10 months), and at three constant temperatures (12, 16, 19 °C) in Puerto Montt (41°S), Central Southern Chile (09/2006-02/2007; 5 months). Female crabs uniformly produced up to 3 viable egg-masses within 4 1/2 months in Antofagasta and in Puerto Montt (at 19 °C). The second egg-mass was observed 62.5 days (± 7.6; N = 7) after the oviposition of the first clutch and a third egg-mass followed 73.5 days (± 12.5; N = 11) later in Antofagasta (at 16-23 °C). Comparably, a second oviposition took place 64.4 days (± 9.8, N = 5) after the first clutch and a third, 67.0 days (± 2.8, N = 2), thereafter, at 19 °C in Puerto Montt. At the two lower temperatures (16 and 12 °C) in Puerto Montt a second egg-mass was extruded after 82.8 days (± 28.9; N = 4) and 137 days (N = 1), respectively. The duration of egg-development from oviposition until larval hatching decreased from 65 days at 12.5 °C to 22.7 days at the observed upper temperature threshold of 22 °C. Based on the derived relationship between temperature and the duration of egg-development (y = 239.3175e^{− 0.107x}; N = 21, r² = 0.83) and data on monthly percentages of ovigerous females from field studies, the annual number of egg-masses of C. setosus was calculated. This analysis revealed an annual output of about one egg-mass close to the species northern and southern distributional limits in Casma (9°S) and Ancud (43°S), respectively, while at Coquimbo (29°S) about two and in Concepción (36°S) more than 3 egg-masses are produced per year.     

Preliminary studies on cryopreservation of snakehead (Channa striata) embryos

This paper reports the findings of the ongoing studies on cryopreservation of the snakehead, Channa striata embryos. The specific objective of this study was to collect data on the sensitivity of C. striata embryo hatching rate to low temperatures at two different developmental stages in the presence of four different cryoprotectants. Embryos at morula and heartbeat stages were selected and incubated in 1 M dimethyl sulfoxide (Me₂SO), 1 M ethylene glycol (EG), 1 M methanol (MeOH) and 0.1 M sucrose solutions at different temperatures for a period of time. Embryos were kept at 24 °C (control), 15 °C, 4 °C and −2 °C for 5 min, 1 h and 3 h. Following these treatments, the embryos were then transferred into a 24 °C water bath until hatch to evaluate the hatching rate. The results showed that there was a significant decrease of hatching rate in both developmental stages following exposure to 4 °C and −2 °C at 1 h and 3 h exposure in each treatment. Heartbeat stage was more tolerant against chilling at −2 °C for 3 h exposure in Me₂SO followed by MeOH, sucrose and EG. Further studies will be conducted to find the best method to preserve embryos for long term storage.     

Temperature effects on zoeal morphometric traits and intraspecific variability in the hairy crab Cancer setosus across latitude

Phenotypic plasticity is an important but often ignored ability that enables organisms, within species-specific physiological limits, to respond to gradual or sudden extrinsic changes in their environment. In the marine realm, the early ontogeny of decapod crustaceans is among the best known examples to demonstrate a temperature-dependent phenotypic response. Here, we present morphometric results of larvae of the hairy crab Cancer setosus, the embryonic development of which took place at different temperatures at two different sites (Antofagasta, 23°45′ S; Puerto Montt, 41°44′ S) along the Chilean Coast. Zoea I larvae from Puerto Montt were significantly larger than those from Antofagasta, when considering embryonic development at the same temperature. Larvae from Puerto Montt reared at 12 and 16°C did not differ morphometrically, but sizes of larvae from Antofagasta kept at 16 and 20°C did, being larger at the colder temperature. Zoea II larvae reared in Antofagasta at three temperatures (16, 20, and 24°C) showed the same pattern, with larger larvae at colder temperatures. Furthermore, larvae reared at 24°C, showed deformations, suggesting that 24°C, which coincides with temperatures found during strong EL Niño events, is indicative of the upper larval thermal tolerance limit. C. setosus is exposed to a wide temperature range across its distribution range of about 40° of latitude. Phenotypic plasticity in larval offspring does furthermore enable this species to locally respond to the inter-decadal warming induced by El Niño. Morphological plasticity in this species does support previously reported energetic trade-offs with temperature throughout early ontogeny of this species, indicating that plasticity may be a key to a species’ success to occupy a wide distribution range and/or to thrive under highly variable habitat conditions.     

Parental size and environmental conditions affect egg capsule production by Nassarius reticulatus (Linnaeus 1758) (Gastropoda: Nassariidae)

In shallow coastal habitats scavenging netted whelks Nassarius reticulatus attached egg capsules to the stipes of red algae Chondrus crispus and occasionally on Furcellaria lumbricalis and Plumaria plumose. In the laboratory egg capsules were laid on aquaria sides and lids by individuals ≥ 21 mm shell length. Larger size classes produced more egg capsules and spawned over a longer period and in doing so partitioned less energy into shell growth. Large netted whelks (25-28.9 mm) produced larger capsules which contained significantly more and larger eggs than those produced by smaller individuals (21-24.9 mm). Egg capsule production continued throughout the year by regularly fed N. reticulatus held at ambient seawater temperatures. Egg production increased in the spring and summer with peak production during June (15 °C), decreased between August and October and resumed again during the winter (November to February at ∼ 7 °C). During the summer (15-16 °C) egg capsules were smaller and contained smaller eggs than those deposited during the winter (7-10 °C), although the number of eggs · capsule⁻¹ was similar. Enforced food limitation reduced the number and size of the egg capsules, the number and size of eggs produced · female⁻¹ and the duration of the breeding period. Hatching success of N. reticulatus egg capsules was high (95%) even at winter seawater temperatures (11-8.5 °C) and the duration of embryonic development was fastest between 15 and 17.5 °C.     

Mycosporine-like amino acids in azooxanthellate and zooxanthellate early developmental stages of the soft coral Heteroxenia fuscescens

The ontogenetic changes of MAAs in the soft coral Heteroxenia fuscescens was studied in relation to their symbiotic state (azooxanthellate vs. zooxanthellate) under different temperature conditions in the Gulf of Eilat, northern Red Sea. The HPLC chromatograms for extracts of the planulae, azoo- and zooxanthellate primary polyps of H. fuscescens from all dates of collection yielded a single peak at 320 nm that has been identified as the compound palythine. Concentration of palythine in planulae at 23 °C was 7.57 ± 1 nmol mg^− 1 protein and at 28 °C reached 17.29 ± 1 nmol × mg^− 1 protein. Concentration of palythine in azooxanthellate primary polyps was 16.4 ± 3 nmol × mg^− 1 protein and 28.37 ± 2.8 nmol × mg^− 1 protein at 23 °C and 28 °C respectively. The palythine concentration for zooxanthellate primary polyps at 23 °C was 13 ± 3 nmol × mg^− 1 protein and at 28 °C 32.7 ± 2 nmol mg^− 1 protein. Palythine concentrations were significantly higher at 28 °C in the different animal groups and correlated linearly with the ambient collection temperature. This study shows for the first time that UVR and temperature act synergistically and affect the MAA levels of early life-history stages of soft corals.     

Temperature-dependent activity in early life stages of the stone crab Paralomis granulosa (Decapoda, Anomura, Lithodidae): A role for ionic and magnesium regulation?

Marine brachyuran and anomuran crustaceans are completely absent from the extremely cold (− 1.8 °C) Antarctic continental shelf, but caridean shrimps are abundant. This has at least partly been attributed to low capacities for magnesium excretion in brachyuran and anomuran lithodid crabs ([Mg²⁺]_HL = 20-50 mmol L^− 1) compared to caridean shrimp species ([Mg²⁺]_HL = 5-12 mmol L^− 1). Magnesium has an anaesthetizing effect and reduces cold tolerance and activity of adult brachyuran crabs. We investigated whether the capacity for magnesium regulation is a factor that influences temperature-dependent activity of early ontogenetic stages of the Sub-Antarctic lithodid crab Paralomis granulosa. Ion composition (Na⁺, Mg²⁺, Ca²⁺, Cl⁻, SO₄²⁻) was measured in haemolymph withdrawn from larval stages, the first and second juvenile instars (crabs I and II) and adult males and females. Magnesium excretion improved during ontogeny, but haemolymph sulphate concentration was lowest in the zoeal stages. Neither haemolymph magnesium concentrations nor Ca²⁺:Mg²⁺ ratios paralleled activity levels of the life stages. Long-term (3 week) cold exposure of crab I to 1 °C caused a significant rise of haemolymph sulphate concentration and a decrease in magnesium and calcium concentrations compared to control temperature (9 °C). Spontaneous swimming activity of the zoeal stages was determined at 1, 4 and 9 °C in natural sea water (NSW, [Mg²⁺] = 51 mmol L^− 1) and in sea water enriched with magnesium (NSW + Mg²⁺, [Mg²⁺] = 97 mmol L^− 1). It declined significantly with temperature but only insignificantly with increased magnesium concentration. Spontaneous velocities were low, reflecting the demersal life style of the zoeae. Heart rate, scaphognathite beat rate and forced swimming activity (maxilliped beat rate, zoea I) or antennule beat rate (crab I) were investigated in response to acute temperature change (9, 6, 3, 1, − 1 °C) in NSW or NSW + Mg²⁺. High magnesium concentration reduced heart rates in both stages. The temperature-frequency curve of the maxilliped beat (maximum: 9.6 beats s^− 1 at 6.6 °C in NSW) of zoea I was depressed and shifted towards warmer temperatures by 2 °C in NSW + Mg²⁺, but antennule beat rate of crab I was not affected. Magnesium may therefore influence cold tolerance of highly active larvae, but it remains questionable whether the slow-moving lithodid crabs with demersal larvae would benefit from an enhanced magnesium excretion in nature.     

Synergistic effects of high temperature and sulfide on tropical seagrass

To examine the synergism of high temperature and sulfide on two dominant tropical seagrass species, a large-scale mesocosm experiment was conducted in which sulfide accumulation rates (SAR) were increased by adding labile carbon (glucose) to intact seagrass sediment cores across a range of temperatures. During the initial 10 d of the 38 d experiment, porewater SAR in cores increased 2- to 3-fold from 44 and 136 μmol L^− 1 d^− 1 at 28-29 °C to 80 and 308 μmol L^− 1 d^− 1 at 34-35 °C in Halodule wrightii and Thalassia testudinum cores, respectively. Labile C additions to the sediment resulted in SAR of 443 and 601 μmol L^− 1 d^− 1 at 28-29 °C and 758 to 1,557 μmol L^− 1 d^− 1 at 34-35 °C in H. wrightii and T. testudinum cores, respectively. Both T. testudinum and H. wrightii were highly thermal tolerant, demonstrating their tropical affinities and potential to adapt to high temperatures. While plants survived the 38 d temperature treatments, there was a clear thermal threshold above 33 °C where T. testudinum growth declined and leaf quantum efficiencies (Fv/Fm) fell below 0.7. At this threshold temperature, H. wrightii maintained shoot densities and leaf quantum efficiencies. Although H. wrightii showed a greater tolerance to high temperature, T. testudinum had a greater capacity to sustain biomass and short shoots under thermal stress with labile C enrichment, regardless of the fact that sulfide levels in the T. testudinum cores were 2 times higher than in the H. wrightii cores. Tropical seagrass tolerance to elevated temperatures, predicted in the future with global warming, should be considered in the context of the sediment-plant complex which incorporates the synergism of plant physiological responses and shifts in sulfur biogeochemistry leading to increased plant exposure to sulfides, a known toxin.     

Effects of temperature, salinity, and irradiance on the growth of the dinoflagellate Akashiwo sanguinea

The effects of temperature, salinity, and irradiance on the growth of the dinoflagellate Akashiwo sanguinea were examined in the laboratory. The irradiance at the light compensation point (I₀) was 14.40 μmol m^− 2 s^− 1 and the irradiance at growth saturation (I_s) was 114 μmol m^− 2 s^− 1. We exposed A. sanguinea to 48 combinations of temperature (5-30 °C) and salinity (5-40) under saturating irradiance; it exhibited its maximum growth rate of 1.13 divisions/day at a combination of 25 °C and salinity of 20. A. sanguinea was able to grow at temperatures from 10 to 30 °C and salinities from 10 to 40. This study revealed that A. sanguinea was a eurythermal and euryhaline organism; in Japan it should have formed blooms in early summer, when salinity was relatively low. In addition, it was noteworthy that A. sanguinea had markedly cold-durability, retaining the motile form of vegetative cells for more than 50 days at 5 °C and at salinities of 25-30.     

Egg production and early development of Thysanoessa inermis and Euphausia pacifica (Crustacea: Euphausiacea) in the northern Gulf of Alaska

Early life history patterns were studied in the dominant euphausiids from the northern Gulf of Alaska (GOA) in 2001-2004. Gravid females of Thysanoessa inermis were observed in April and May. Brood size varied from 10 to 1021 eggs with an average of 138 ± 19 (95% CI) eggs female^− 1. Most gravid females started to release eggs within the first 2 days of incubation. The average number of eggs released per female was similar in incubation Day 1 and 2, but significantly smaller on Day 3 and 4. About 25% of the females were continuously releasing eggs over 3 days rather than producing a single distinctive brood. In contrast, gravid females of Euphausia pacifica were observed from early July through October. Most gravid females released eggs on the first day of observation, while only 2% of females produced eggs repeatedly. Brood size varied from 20 to 246 eggs with an average of 102 ± 12 (95% CI) eggs female^− 1. The relationship between E. pacifica brood size and ambient chlorophyll-a concentration was sigmoidal (r² = 0.73), with food saturated brood size of 144 ± 14(SE, P < 0.001) eggs, and half-saturation occurring at 0.46 ± 0.02(SE, P < 0.001) mg chlorophyll-a m^− 3. The average interbrood interval of E. pacifica reared at 12 °C and satiated food conditions in the laboratory was ∼ 8 days, suggesting their potential individual fecundity in the GOA was 1148-1530 eggs per spawning season. Hatching and early development (from egg to furcilia stage) was studied under 5 °C, 8 °C and 12 °C. Hatching was nearly synchronous and lasted 3-6 h, depending on incubation temperature. Development times from egg to the first furcilia stage ranged between 20 and 33 days for T. inermis, and 15 and 45 days for E. pacifica at 12 °C and 5 °C, respectively.     

Seed germination of Lasia spinosa as a function of temperature,light, desiccation,and storage

Lasia spinosa seeds were not dormant at maturity in early spring. The most favorable temperatures for germination were between 25 and 30 °C, and final percentage and rate of germination decreased with an increase or decrease in temperature. When L. spinosa seeds were transferred to 25 °C, after 60 days at 10 °C (where none of the seeds germinated), final germination increased from 0% to 78%. Seeds germinated to high percentage both in light and in dark, although dark germination took more than twice as long as in the light. During desiccation of seeds at 15 °C and 45% relatively humidity, moisture loss decreased exponentially from 2.02 to 0.13 g H₂O g⁻¹ dry wt within 16 days, and only a few seeds (12%) survived 0.13 g H₂O g⁻¹ dry wt moisture content. Seeds stored at 0.58 g H₂O g⁻¹ dry wt moisture content at four constant temperatures (4, 10, 15, and −18 °C) for up to 6 months exhibited a well-defined trend of decreasing viability with decreasing temperature. Thus, we concluded that freshly harvested L. spinosa seeds are non-dormant and recalcitrant. Also, the seeds with 0.58 g H₂O g⁻¹ dry wt moisture content could be effectively stored for a few months between 10 and 15 °C although the most appropriate temperature for wet storage appears to be 10 °C, as it is close to the minimum temperature for germination and so there will be less pre-sprouting compared to 15 °C.     

Changes in zoeae of the snow crab, Chionoecetes opilio, with variation in incubation temperature

The purpose of this study was to evaluate the effect of embryonic incubation temperature on the morphology, energetic content, and individual weights of C. opilio zoeae. Ovigerous female C. opilio were collected by trawl from the eastern Bering Sea in the summer of 2002 and were held in the laboratory at five temperature treatments (− 1, 0, 1, 3, and 6 °C) from embryo stage five to hatching. The mean duration of incubation ranged from 240 d at 6 °C to 353 d at − 1 °C. No relationship between larval weight and caloric content with incubation temperature was detected, indicating that increased development time at cooler temperatures may not have an energetic cost. However, several morphological features varied with temperature. Most larval characters including rostro-dorsal length, rostral spine length, lateral spine length, and carapace width were smaller at the warmest incubation temperature. Protopodite length did not differ with temperature. The length of the 3rd abdominal somite consistently increased with increased temperature and may serve as a suitable indicator that could be used in field populations to infer temperature of incubation. Variation in morphology with temperature could confound identification of C. opilio zoeae from those of a congener, Chionoecetes bairdi because identification of C. bairdi is based on the length of the 3rd abdominal somite.     

Laboratory experiments on the effects of variable suspended sediment concentrations on the ecophysiology of the porcelain crab Petrolisthes elongatus (Milne Edwards, 1837)

The porcelain crab Petrolisthes elongatus is a particulate suspension feeding species common to coastal areas of New Zealand (NZ). Consistent with the responses of other suspension feeding species, it is likely to be negatively influenced by elevated suspended sediment concentrations. Laboratory experiments were conducted to quantify the effect of temperature (12 °C, 15 °C and 18 °C) and suspended sediment concentration (total particulate matter (TPM): low < 100 mg L^− 1; medium 100-1000 mg L^− 1; high > 1000 mg L^− 1) on the clearance rate (CR in L h^− 1), oxygen uptake rate (VO₂ in mL h⁻¹), net absorption efficiency (AE), and net energy budget (NEB in J h^− 1) of P. elongatus across a range of sizes. Variation in CR and AE was independent of temperature and of body size, but were significantly different (P < 0.05) at low and medium suspended sediment concentrations compared with high suspended sediment concentrations. CR responded in a non-linear manner to changes in TPM, increasing with TPM up to a maximum value at medium-low concentrations (approximately 250 mg L^− 1) and then decreasing thereafter. CR had almost completely shut down at TPM concentrations of > 1000 mg L^− 1 and at particulate organic matter (POM) concentrations of > 250 mg L^− 1. AE was zero at approximate TPM and POM values of 1200 mg L^− 1 and 300 mg L^− 1, respectively. VO₂ was positively correlated with body size and with temperature, but was independent of TPM. NEB values for P. elongatus were low (approx 110 J g^− 1 h^− 1) at low sediment concentrations, were high (approx 320 J g^− 1 h^− 1) at medium sediment concentrations, and were negative (approx − 114 J g^− 1 h^− 1) at high sediment concentrations. These findings indicate that P. elongatus is likely to be food-limited at sediment concentrations of < 100 mg L^− 1, and severely negatively affected at sediment concentrations of > 1000 mg L^− 1, at least for the duration of such events which may persist for 2-3 days in coastal environments where this crab occurs.     

Rapid down-regulation of mitochondrial fat metabolism in human muscle after training cessation is dissociated from changes in insulin sensitivity

The association between impairment in mitochondrial muscle fat oxidative capacity (OX_FA) and occurrence of insulin resistance was examined in 14 healthy trained men (age, 24 ± 4 yr) submitted to 4 weeks of training cessation. Training stop induced a significant decrease in mRNA levels of proteins involved in muscle fat metabolism, particularly PPARα (−58%, P < 0.01) and PGC-1α (−30%, P < 0.05), a 21% reduction in OX_FA (P < 0.01), and reduced fat oxidation during moderately intense exercise (P < 0.05). In contrast, there was no significant alteration in insulin sensitivity. In conclusion, decline in OX_FA is a rapid metabolic event following training cessation. It is involved in the regulation of whole body fat balance but not in the deterioration of insulin sensitivity.     

Rapid thermal responses and thermal tolerance in adult codling moth Cydia pomonella (Lepidoptera: Tortricidae)

In order to preserve key activities or improve survival, insects facing variable and unfavourable thermal environments may employ physiological adjustments on a daily basis. Here, we investigate the survival of laboratory-reared adult Cydia pomonella at high or low temperatures and their responses to pre-treatments at sub-lethal temperatures over short time-scales. We also determined critical thermal limits (CTLs) of activity of C. pomonella and the effect of different rates of cooling or heating on CTLs to complement the survival assays. Temperature and duration of exposure significantly affected adult C. pomonella survival with more extreme temperatures and/or longer durations proving to be more lethal. Lethal temperatures, explored between −20 °C to −5 °C and 32 °C to 47 °C over 0.5, 1, 2, 3 and 4 h exposures, for 50% of the population of adult C. pomonella were −12 °C for 2 h and 44 °C for 2 h. Investigation of rapid thermal responses (i.e. hardening) found limited low temperature responses but more pronounced high temperature responses. For example, C. pomonella pre-treated for 2 h at 5 °C improved survival at −9 °C for 2 h from 50% to 90% (p < 0.001). At high temperatures, pre-treatment at 37 °C for 1 h markedly improved survival at 43 °C for 2 h from 20% to 90% (p < 0.0001). We also examined cross-tolerance of thermal stressors. Here, low temperature pre-treatments did not improve high temperature survival, while high temperature pre-treatment (37 °C for 1 h) significantly improved low temperature survival (−9 °C for 2 h). Inducible cross-tolerance implicates a heat shock protein response. Critical thermal minima (CTmin) were not significantly affected by cooling at rates of 0.06, 0.12 and 0.25 °C min⁻¹ (CTmin range: 0.3-1.3 °C). By contrast, critical thermal maxima (CTmax) were significantly affected by heating at these rates and ranged from 42.5 to 44.9 °C. In sum, these results suggest pronounced plasticity of acute high temperature tolerance in adult C. pomonella, but limited acute low temperature responses. We discuss these results in the context of local agroecosystem microclimate recordings. These responses are significant to pest control programmes presently underway and have implications for understanding the evolution of thermal tolerance in these and other insects.     

Near anoxia and sulfide as possible factors influencing the spatial distribution of Acartia tonsa and Acartia clausi: Comparative evaluation of egg tolerance

In many coastal and estuarine areas the planktonic copepods Acartia tonsa and Acartia clausi show a spatial separation with A. tonsa restricted to brackish waters and confined environments and A. clausi inhabiting areas more influenced by sea water. The hatching and viability of A. tonsa and A. clausi eggs exposed to anoxia and anoxia/sulfide (conditions that are frequent in bottom waters of the most confined areas) was evaluated to determine if these stress factors play a role in the distribution of these species. Subitaneous eggs, spawned by laboratory reared organisms, were incubated in near anoxia (< 7.59 × 10^− 3 mmol O₂ L^− 1) or anoxia/sulfide (∼ 1 mmol L^− 1) for different periods (1, 4, 8 and 15 days), then transferred to normoxic conditions. The exposure of the eggs to near anoxia and sulfide appears to induce the same response (quiescence) in both species. Exposure times ≤ 8 days to near anoxia or anoxia/sulfide did not affect egg viability, while 15 day exposure caused significant declines in hatching success of both species. A significant difference between the effects of near anoxia and anoxia/sulfide was observed when incubation lasted 15 days; hatching of eggs exposed to sulfide being higher than that of eggs exposed to near anoxia for both species. No significant differences were observed between the two species in hatching success of eggs exposed to both near anoxia and anoxia/sulfide (with the exception of eggs incubated in near anoxia for 4 days). The results indicate that the impact of anoxia and sulfide on the eggs of the two Acartia species cannot be a factor explaining the spatial distribution in coastal and brackish environments of these copepods.Feeding experiments on A. clausi were also performed. Suitability of different algal species to rear this copepod was evaluated and the results were compared with data previously obtained for A. tonsa. Differences in feeding needs between A. clausi and A. tonsa are discussed.     

The timing of leaf fall affects cold acclimation by interactions with air temperature through water and carbohydrate contents

Three parameters (i.e. the water content, soluble sugar content and minimal air temperature) can be used to predict the cold acclimation process of walnut trees. In order to test this assumption, two-year-old walnuts were defoliated at two different dates, i.e. mechanical defoliation in early October (early leaf fall, EF) or natural defoliation in early November (natural leaf fall, NF) and conditioned in either outdoor freeze-deprived or cold-deprived (T_min > 13 °C) greenhouses over winter. Even if early defoliation date could have affected short day signal perception (SDSP), water balance and carbohydrate metabolism were more altered. EF treatment, by stopping transpiration, significantly increased tree's water content and at warm temperature high root activity stopped normal winter dehydration. Starch content decreased in all treatments, but there was only a significant increase in soluble sugar content when water content had sufficiently decreased. Thus, depending on date of defoliation, cold-deprived trees were or were not able to acclimate to frost (minimal frost hardiness = −21.8 °C vs. −22.1 °C in controls (freeze-deprived) for NF and −13.7 °C vs. −25.3 °C in controls for EF). Different treatments showed the relationship between minimal water content observed during winter and maximal soluble sugars synthesized. Thus, the cold acclimation process appeared dependent on these physiological parameters (water and soluble sugar contents) through the interaction between air temperature and timing of leaf fall.     

Metabolic thermogenesis and evaporative water loss in the Hwamei Garrulax canorus

Basal metabolic rate (BMR) is thought to be a major hub in the network of physiological mechanisms connecting life history traits. Evaporative water loss (EWL) is a physiological indicator that is widely used to measure water relations in inter- or intraspecific studies of birds in different environments. In this study, we examined the physiological responses of summer-acclimatized Hwamei Garrulax canorus to temperature by measuring their body temperature (T_b), metabolic rate (MR) and EWL at ambient temperatures (T_a) between 5 and 40 °C. Overall, we found that mean body temperature was 42.4 °C and average minimum thermal conductance (C) was 0.15 ml O₂ g⁻¹ h⁻¹ °C⁻¹ measured between 5 and 20 °C. The thermal neutral zone (TNZ) was 31.8–35.3 °C and BMR was 181.83 ml O₂ h⁻¹. Below the lower critical temperature, MR increased linearly with decreasing T_a according to the relationship: MR (ml O₂ h⁻¹)=266.59–2.66 T_a. At T_as above the upper critical temperature, MR increased with T_a according to the relationship: MR (ml O₂ h⁻¹)=−271.26+12.85 T_a. EWL increased with T_a according to the relationship: EWL (mg H₂O h⁻¹)=−19.16+12.64 T_a and exceeded metabolic water production at T_a>14.0 °C. The high T_b and thermal conductance, low BMR, narrow TNZ, and high evaporative water production/metabolic water production (EWP/MWP) ratio in the Hwamei are consistent with the idea that this species is adapted to warm, mesic climates, where metabolic thermogenesis and water conservation are not strong selective pressures.     

Preservation at ultra-low temperature of in vitro cultured arbuscular mycorrhizal fungi via encapsulation–drying

At present, over 300 species of arbuscular mycorrhizal fungi (AMF) have been identified, most of which being stored in international collections. Their maintenance is mostly achieved in greenhouse via continuous culture on trap plants or in vitro in association with excised root organs. Both methods are work-intensive and for the former present the risk of unwanted contaminations. The in vitro root organ culture of AMF has become an alternative preventing contamination. Nevertheless, the risk for somaclonal variation during the sub-cultivation process cannot be excluded. A method for the long-term conservation that guarantees the stability of the biological material is thus highly demanded to preserve the microorganisms and their genetic stability. Here, 12 AMF isolates cultured in vitro in association with excised carrot roots were encapsulated in alginate beads and subsequently cryopreserved. Several protocols were tested taking into consideration culture age, alginate bead pre-drying, and rate of decrease in temperature. The viability of the AMF isolates was estimated by the percentage of potentially infective beads (%PIB) that measure the % of beads that contain at least one germinated propagule. Thermal behaviour of alginate beads was analysed by a differential thermal calorimeter before and after drying to estimate the frozen and unfrozen water during the cryopreservation process. It was shown that the spore damage was directly related to ice formation during cryopreservation. The encapsulation and culture age were also determinant parameters for the successful cryopreservation. Irrespective of the AMF isolate, the optimal procedure for cryopreservation comprised five steps: (1) the encapsulation of propagules (i.e. spores and mycorrhizal root pieces) isolated from 5 m old cultures, (2) the incubation overnight in trehalose (0.5 M), (3) the drying during 48 h at 27 °C, (4) the cryopreservation in the freezer at −130 °C following a two-step decrease in temperature: a fast decrease (∼12 °C min⁻¹) from room temperature (+20 °C) to −110 °C followed by a slow decrease in temperature (∼1 °C min⁻¹) from −110 °C to −130 °C, and (5) the direct thawing in a water bath (+35 °C). The % PIB was above 70 % for all the isolates and even above 95 % for 11 out of the 12 isolates after several months of storage at ultra-low temperature. All the isolates kept their capacity to associate to an excised carrot root in vitro and to reproduce the fungal life cycle with the production of several hundreds to thousands of spores after 2 m. This method opens the door for the long-term maintenance at ultra-low temperature of AMF isolates within international repositories.     

Determination of thermal properties of composting bulking materials

Thermal properties of compost bulking materials affect temperature and biodegradation during the composting process. Well determined thermal properties of compost feedstocks will therefore contribute to practical thermodynamic approaches. Thermal conductivity, thermal diffusivity, and volumetric heat capacity of 12 compost bulking materials were determined in this study. Thermal properties were determined at varying bulk densities (1, 1.3, 1.7, 2.5, and 5 times uncompacted bulk density), particle sizes (ground and bulk), and water contents (0, 20, 50, 80% of water holding capacity and saturated condition). For the water content at 80% of water holding capacity, saw dust, soil compost blend, beef manure, and turkey litter showed the highest thermal conductivity (K) and volumetric heat capacity (C) (K: 0.12–0.81 W/m °C and C: 1.36–4.08 MJ/m³ °C). Silage showed medium values at the same water content (K: 0.09–0.47 W/m °C and C: 0.93–3.09 MJ/m³ °C). Wheat straw, oat straw, soybean straw, cornstalks, alfalfa hay, and wood shavings produced the lowest K and C values (K: 0.03–0.30 W/m °C and C: 0.26–3.45 MJ/m³ °C). Thermal conductivity and volumetric heat capacity showed a linear relationship with moisture content and bulk density, while thermal diffusivity showed a nonlinear relationship. Since the water, air, and solid materials have their own specific thermal property values, thermal properties of compost bulking materials vary with the rate of those three components by changing water content, bulk density, and particle size. The degree of saturation was used to represent the interaction between volumes of water, air, and solids under the various combinations of moisture content, bulk density, and particle size. The first order regression models developed in this paper represent the relationship between degree of saturation and volumetric heat capacity (r = 0.95–0.99) and thermal conductivity (r = 0.84–0.99) well. Improved knowledge of the thermal properties of compost bulking materials can contribute to improved thermodynamic modeling and heat management of composting processes.     

Effect of desiccation level and storage temperature on green spore viability of Osmunda japonica

In order to effectively preserve green spores, which have relatively higher water content and lose viability more quickly than non-green spores, we studied the effect of desiccation level and storage temperature on Osmunda japonica spores. The water content of fresh spores was 11.20%. After 12 h desiccation by silica gel, the water content decreased to 6% but spore viability did not change significantly. As the desiccation continued, the decrease in water content slowed, but spore viability dropped. For almost all storage periods, the effects of storage temperature, desiccation level, and temperature × desiccation level were significantly different. After seven days of storage, spores at any desiccation level stored at 4 °C obtained high germination rates. After more than seven days storage, liquid nitrogen (LN) storage obtained the best results. Storage at −18 °C led to the lowest germination rates. Spores stored at room temperature and −18 °C all died within three months. For storage at 4 °C and in LN, spores desiccated 12 and 36 h obtained better results. Spores without desiccation had the highest germination rates after being stored at room temperature, but suffered the greatest loss after storage at −18 °C. These results suggest that LN storage is the best method of long-term storage of O. japonica spores. The critical water content of O. japonica spores is about 6% and reduction of the water content to this level improves outcome after LN storage greatly. The reason for various responses of O. japonica spores to desiccation and storage temperatures are discussed.