Induction of developmental abnormalities in larval goldfish, Carassius auratus L., under cool incubation conditions

Compared with incubation at a constant 22° C, exposure of goldfish embryos and larvae to 13° C, under a variety of thermal protocols, caused increased frequencies of abnormal development and, in some cases, reduced survival to hatching. The low-temperature incubation conditions were particularly deleterious when eggs were incubated at 13° C from the outset, regardless of the temperature at which the donor female ovulated and the eggs were fertilized. Significantly higher frequencies of developmental abnormalities were also noted when embryos were transferred from 22°C to 13°C at 6, 24, 128 and, in one case, 175 h after fertilization. In three of five experiments, subjecting embryos and larvae to diel fluctuations between 22 and 13° C, with a 5-h hold at the lower temperature, caused an increase in development abnormalities. These results demonstrate that the thermal requirements of goldfish embryos and larvae necessitate a delay in ovulation and spawning until water is sufficiently warm. Developmental abnormalities can be induced by exposure to cool (13° C) conditions, at least up to the time that swimbladder inflation occurs.     

Toxicity of crude extracellular products of Aeromonas hydrophila in tilapia, Tilapianilotica

Extracellular products (ECP) secreted from Aeromonas hydrophila with haemolytic andproteolytic activity were studied with respect to temperature and time of incubation as well as thelethal toxicity on tilapia, Tilapia nilotica . The highest production of the haemolysin productwas achieved when Aer. hydrophila was grown at 35°C for 30 h. Tilapia erythrocytewas found to be more susceptible than sheep erythrocyte for determining the haemolytic activity.The haemolytic activity against tilapia erythrocyte was completely inactivated after heating theECP at 60°C for 10 min or 55°C for 15 min. The proteolytic activity was maximized whenthe bacterium was grown at 30°C for 36 h. Complete inactivation of the protease enzyme wasperformed after heating the ECP at 80°C for 10 min or 70°C for 15 min. Aeromonashydrophila was found to produce haemolytic and proteolytic exotoxin lethal to tilapia (LD₅₀ 2·1 × 10⁴ cell/fish), as well as heat stable unknown virulent factors thatwere responsible for 20% mortality. The lethality of ECP was decreased by heating andcompletely inactivated by boiling at 100°C for 10 min.     

Relationships between maternal size, egg diameter, time of spawning season, temperature, and length at hatch of Atlantic silverside, Menidia menidia

Eggs were stripped from gravid Atlantic silversides collected on two occasions, once during the early part and once during the late part of the natural spawning season. Unfertilized egg diameter was not correlated with length of the female, nor was it significantly larger during the early part of the season. Eggs were fertilized and incubated in the laboratory. Larval length at hatch was measured every 24 h during the hatching period after embryos were incubated at 18 or 25° C. Lower incubation temperature caused a significantly greater length at hatch for the offspring of each of the 20 females studies. In most cases (17 out of 20 at 25° C, 10 out of 20 at 18° C), there was a significant decrease in length at hatch during the hatching period for a given female's eggs incubated at a given temperature. In the natural environment, larvae hatched early in the season under cooler temperatures could average 12% longer than those hatched later under warmer temperatures, and therefore may have a greater chance of survival. The results help to explain the observation that field-caught M. menidia that hatched early in the season are larger at any given age than those that hatched late in the season.     

Elevation of the heat resistance of Salmonella typhimurium by sublethal heat shock

The survival of Salmonella typhimurium after a standard heat challenge at 55°C for 25 min increased by several orders of magnitude when cells grown at 37°C were pre-incubated at 42°, 45° or 48°C before heating at the higher temperature. Heat resistance increased rapidly after the temperature shift, reaching near maximum levels within 30 min. Elevated heat resistance persisted for at least 10 h. Preincubation of cells at 48°C for 30 min increased their resistance to subsequent heating at 50°, 52°, 55°, 57° or 59°C. Survival curves of resistant cells were curvilinear. Estimated times for a '7D' inactivation increased by 2.6- to 20-fold compared with cells not pre-incubated before heat challenge.     

Temperature-dependent development rates of cod Gadus morhua eggs

Temperature development relationships were determined for batches of Irish Sea cod Gadus morhua eggs incubated in flow-through incubators. Hatching began 16·4 days after fertilization (DAF) at 6° C, 10·3 DAF at 8° C, 9·4 DAF at 10° C and 7·4 DAF at 12° C. Egg mortality increased at the higher temperatures, but survival was >80%. Results were compared with published data at four comparable stage end points: the end of blastula, the end of gastrula, the point of growth of the embryo completely surrounding the yolk and the point when 50% of the eggs were hatched. All the studies showed a curvilinear relationship between age at stage and temperature. There was a 12 day inter-study difference in time to 50% hatch at 2° C and 4 day difference at 10° C. There were no consistent trends that differentiated eastern v. western, or northern v. southern populations. A single model for cod egg incubation time from fertilization to 50% hatch was derived based on data from six cod populations, but it is recommended that individual stock relationships should be used where possible.     

Incubation of eggs of tuatara, Sphenodon punctatus

Eggs of the tuatara, Sphenodon punctatus , were incubated either buried or half buried in vermiculite at constant temperatures of 15, 18, 20, 22 and 25 °C and constant water potentials between —90 and —400 kPa. Many clutches failed completely, possibly because they had been taken from females prior to proper shell development. Failed eggs were significantly smaller than successful eggs. Incubation is unsuccessful at 15 °C. Hatching success is high between 18 and 22 °C but low at 25 °C, but equally successful between 18 and 22°C. Incubation is strongly influenced by temperature, with mean incubation periods of 328 days at 18 °C, 259 days at 20 °C, 169 days at 22 °C and 150 days at 25 °C. Water potential generally has little influence on incubation time at a given temperature. Buried eggs hatch sooner than partially buried eggs at 20 °C but the large range makes significance dubious.
Eggs on the driest substrata at 18 and 20 °C lose water initially but then gain water through the rest of incubation. Eggs in all other conditions gain water throughout incubation, with the rate of i water absorption being maintained or increasing late in incubation. The suggestion that increasing rate of water absorption late in incubation facilitates explosive hatching is not supported. Egg mass at the time of hatching varies from 132 to 398% of initial values, depending on incubation conditions. Final egg mass is not affected significantly by incubation temperature. Hence, rates of absorption increase with temperature.
Water potential has no influence on hatchling size. However, hatchlings from buried eggs generally are significantly larger than those from partially buried eggs.     

Reversible change in embryonic diapause intensity by mild temperature in the Chinese rice grasshopper, Oxya chinensis

The effects of temperature on maintenance and termination of embryonic diapause were investigated in Jining (35.4°N, 116.6°E) and Sihong (33.5°N, 118.2°E) strains of the Chinese rice grasshopper, Oxya chinensis Thunberg (Orthoptera: Catantopidae). Eggs of both strains entered diapause when incubated at 30, 25, or 20 °C. Chilling at 8 °C had an evident effect on diapause termination and almost all eggs chilled for 60 days ended diapause development. Chilling of eggs at 8 °C for only 20 days failed to result in any hatching at 20 °C, suggesting that such level of chilling was not enough to induce diapause termination. However, the treatment combining incubation of eggs at 30 °C for varying lengths of time with subsequent incubation to 20 °C had a distinct effect on the completion of diapause of the eggs. The results indicate that there were two temperature optima, that is, low temperature (chilling) and high temperature, for diapause development in this grasshopper species. Incubation of chilled eggs at 20 °C for 5–15 days followed by further incubation at 25 °C reduced termination of diapause significantly compared with the eggs only chilled at 8 °C. Exposure of eggs chilled at 8 °C to a pulse of 25 °C from 1 to 7 days, separated by a 20-day interval at 8 °C, resulted in a decrease in the percentage of successfully hatched eggs as the length of the pulse of 25 °C increased. The results suggest that diapause intensity may be restored at moderately high temperatures. This reversible change in diapause intensity would play an important role in maintaining diapause before winter.     

The embryonic development and the thermal effects on the development of the mountain whitefish, Prosopium williamsoni (Girard)

The general development of the mountain whitefish was similar to other whitefishes. Mountain whitefish required more thermal units to reach corresponding stages of the lake whitefish, after the stage when the blastodisc was prominently raised on the yolk. The development process was greatly disrupted and a heavy mortality of eggs occurred when the temperature was raised to 9° C or higher. In hatched larvae abnormalities like monomicrophthalmia, agape jaws, and coloboma occurred when the temperature was raised to 9° C or higher, although the reported optimum temperature for growth of the post yolk sac larvae and juveniles was 9° C-12° C. The late spawned eggs exhibit adaptive mechanisms which enabled them to shorten the incubation time and hatch along with the early spawned eggs, when conditions for survival are at optimum.     

The effects of temperature on the development and mortality of eggs of perch, Perca fluviatilis

SUMMARY. The rate of development and mortality of perch Perca fluviatilis was studied at ten different constant temperatures. The rate of development was inversely related to the incubation temperature, whereas the rate of mortality was directly related to the incubation temperature. The sum of heat (Σ H , degree-days) required for 10, 50 and 90% of the eggs to hatch was found to be constant, regardless of the incubation temperature, with mean values (with 95% confidence limits) of 91.4 (83.3–102.0) degree-days above 4.6°C for 10% hatched, 97.0 (90.9–104.2) degree-days above 4.9°C for 50% hatched and 101.0 (94.3–108.7) degree-days above 5.0°C for 90% hatched. Mortality among the different embryological stages was highest for the pre-hatching stage (i.e. when eye-pigment has been formed) at all temperatures. High mortality among the early stages occurred at temperatures below 8°C and above 12°C.     

Thermal conditioning of fifth-instar Cydia pomonella (Lepidoptera: Tortricidae) affects HSP70 accumulation and insect mortality

Abstract.  Levels of HSP70 protein of fifth-instar codling moth [ Cydia pomonella (L.) (Lepidoptera: Tortricidae)] are determined after conditioning at 35 °C for different times and also after recovery at 22 °C. Protein samples from larvae conditioned for different times are separated by sodium dodecyl sulphate-polyacrylamide gel electrophoresis electrophoresis. Sub-lethal thermal conditioning at 35 °C for 40 min, 2, 6 and 18 h induces new protein bands in the extracts from treated codling moth larvae. Immunodetection with an antibody to a heat-inducible HSP70 indicates a stronger reaction after 35 °C for 2, 6 and 18 h than after 35 °C for 40 min or control and, during the recovery period at 22 °C, the level of heat shock protein decreases. Conditioning of fifth-instar codling moths at 35 °C also induces thermotolerance in the insects and necessitates longer times at a lethal temperature to ensure mortality. Thermotolerance is correlated with the accumulation of heat inducible HSP70 protein.     

Influence of temperature on egg hatching, growth and survival of larvae of Heterobranchus longifilis Val. 1840 (Teleostei: Clariidae)

Eggs of Heterobranchus longifilis Val. 1840 were artificially fertilized and incubated at a range of temperatures (20, 23, 25, 27, 29 and 32°C). The time from fertilization to hatching decreased with increasing temperature. No eggs survived to hatch at 20 and 32°C incubation temperatures, while at 23 and 29°C hatching was only minimal. Optimum hatching was obtained at 25 and 27°C, which corresponds to the ambient temperature range during the breeding season. Larvae of H. longifilis were reared for 11 days post-hatching at 20, 25, 27, 29 and 32°C. Growth increased with temperature (P < 0.05), whereas survival depicted an inverse relationship. Growth was minimal at 20°C and larvae rarely survived to the end of the experiment. Optimum temperature for the primary nursing of H. longifilis larvae was within the 25–27°C temperature range.     

Induction of Fertilization Membrane Formation and Cyanide-insensitive Respiration in Sea Urchin Eggs by the Treatment with Dimethylsulfoxide Followed by an Incubation in an Ice Bath

In unfertilized eggs of the sea urchin Hemicentrotus pulcherrimus , fertilization membrane formation was induced by an incubation with dimethylsulfoxide (DMSO) for several min at 20°c followed by another incubation in an ice bath. The number of eggs with fertilization membrane, thus obtained, increased in relation to the concentration of DMSO between 1 and 3% (v/v) and was higher than 75% at concentrations above 3%. Fertilization membrane formation by this treatment occurred in Ca²⁺ free- or Ca²⁺, Mg²⁺ free- artificial sea water containing EGTA (50 mM) and was inhibited by verapamil. In the presence of DMSO, the membrane formation was also induced by 2, 4-dinitrophenol or cyanide in considerable number of eggs at 20°c. Eggs remained fertilizable, even when they were kept with DMSO for 1 hr at 20°c. DMSO slightly enhanced respiratory rate in unfertilized eggs and substantially reduced it in fertilized eggs. DMSO-treated eggs exhibited cyanide-insensitive respiratory burst following chilling in an ice bath or by adding DNP or cyanide, in a similar manner to the burst induced by sperm.     

Combining heat treatment and subsequent incubation temperature to prevent growth from spores of non-proteolytic Clostridium botulinum

Refrigerated processed foods of extended durability rely on a mild heat treatment combined with refrigerated storage to ensure microbiological safety and quality. The principal microbiological safety risk in foods of this type is non-proteolytic Clostridium botulinum. In this article the combined effect of mild heat treatment and refrigerated storage on the time to growth and probability of growth from spores of non-proteolytic Cl. botulinum is described. Spores of non-proteolytic Cl. botulinum (two strains each of type B, E and F) were heated at 90°C for between 0 and 60 min and subsequently incubated at 5°, 10° or 30°C in PYGS broth in the presence or absence of lysozyme. The number of spores that resulted in turbidity depended on the combination of heat treatment, incubation time and incubation temperature they received. Heating at 90°C for 1 or more min ensured a 10⁶ reduction when spores were subsequently incubated at 5°C for up to 23 weeks. Heating at 90°C for 60 min ensured a 10⁶ reduction over 23 weeks when subsequent incubation was at 10°C in the presence of added lysozyme. The same treatment did not reduce the spore population by 10⁶ when subsequent incubation was at 30°C.     

Fertilization in landlocked sea lamprey: storage of gametes, optimal sperm: egg ratio, and methods of assessing fertilization success

Ovulated, unfertilized eggs of sea lamprey Petromyzon marinus could be stored for 1 day at 15° C without significant loss of fertilizing ability. After 2 days storage most eggs could still be fertilized. Lamprey semen could be stored up to 1 day. Thereafter, a decrease in sperm fertilizing ability occurred, accompanied with a decrease in sperm motility. Unlike teleost fish, sea lamprey eggs could still be fertilized after 1 h contact with water. This extended time of gamete fertility after release into water may help to account for the reproductive success of this species. Maximal fertilization rates were obtained at a sperm: egg ratio of 50 000, a ratio recommended for studies on fertility of individual males. Assessing fertilization success 3 min after fertilization (at cytoplasmic bleb stage) or 5 h after fertilization (at two–cell embryo) was strongly correlated ( r =0·92 and 0·98) with estimation and fertilization success at hatching. These results offer improvement in artificial fertilization techniques under laboratory conditions and provide new information on the biology of fertilization in sea lamprey.     

Temperature-induced changes of survival, development and yolk partitioning in Chondrostoma nasus

Fertilized Chondrostoma nasus eggs were incubated at 10, 13, 16 and 19° C until full resorption of the yolk sac. High survival was observed at 10–16° C (89–92% at the onset of external feeding), whereas at 19) C survival was depressed (76%). The time at which 5, 50 and 95% of individuals had hatched, filled the swim bladder, ingested the first food and fully resorbed the yolk sac was determined. An increase in temperature accelerated development and made it more synchronous. Within the period from fertilization to hatching embryonic development was theoretically arrested (t_{0 dev}) at 8·8° C, and growth was arrested (t_0gr) at 8·86° C. For the whole endogenous feeding period (from fertilization to full yolk resorption) the amount of matter transformed into tissue was temperature independent between 10° and 19° C. Respiration increased exponentially with age; the respiration increase was faster at higher temperatures, but, in general, metabolic expenditures of C. nasus were low. As a consequence, the efficiency of utilizing yolk energy for growth was high as compared with other fish species (57% during the whole endogenous feeding period); it was temperature independent. However, time was used less efficiently at low temperatures, increasing a risk of predation. Within the endogenous feeding period a shift from lower to higher temperatures for optimal yolk utilization efficiency was observed. The temperatures optimal for survival and energetic performance seem to be 13–16° C for egg incubation and 15–18° C for rearing of yolk-feeding larvae. Chondrostoma nasus is a potential candidate for aquaculture for restocking purposes.     

Effect of culture age, pre-incubation at low temperature and pH on the thermal resistance of Aeromonas hydrophila

S. CONDON, M.L. GARCIA, A. OTERO AND F.J. SALA. 1992. The thermal resistance of Aeromonas hydrophila strain NCTC 8049 was determined within the range 48°-65°C with a thermoresistometer TR-SC and McIlvaine buffer. The effects of culture age, pre-incubation at 7°C and the pH of the heating menstruum were evaluated. The pattern of thermal death was dependent on culture age. Cells heated in the late logarithmic growth phase (15 h at 30°C) were twice as resistant as those in the early stage (5 h at 30°C), and the maximum D -value was obtained after 72 h incubation (5.5 total increase). The age of the cells did not affect z -values significantly. The heat resistance of cells incubated for 48 h at 30°C increased (twice) after holding at 7°C for 72 h. Pre-incubation at low temperature of older cultures (72 h, 30°C) did not influence their D -values. Maximum heat resistance was found at pH 6.0 and minimal at pH 4.0. Decreasing the pH from 6.0 4.0 reduced D -values by a factor of 5. Although the strain studied was heat-sensitive ( D _55°C= 0.17 min; z = 5.11°C), survivor curves of cultures older than 50 h showed a significant tailing. Organisms surviving in the tails were only slightly more resistant than were the original population.     

The effect of temperature on the egg incubation period of Capnia bifrons (Plecoptera: Capniidae) from Windermere (English Lake District)

Gravid females of Capnia bifrons (Newman) from Windermere (English Lake District) were almost completely ovoviviparous, the eggs hatching within 15 min after oviposition in the water. When kept in the laboratory at constant temperatures between 3.8 and 19.8°C, few females survived to lay eggs at temperatures above 12.1°C. The relationship between air temperature (T°C) and the egg incubation period (Y days between fertilisation and oviposition) was given by the regression equation: Y = 316.4 T^−0.9996 (r²= 0.957, p < 0.001). This equation successfully predicted egg incubation periods for gravid females kept in cages in the field.
Comparisons with similar studies on four non-ovoviviparous species of Plecoptera showed that egg development was rarely more rapid in C. bifrons . It was also shown that the hypothesis of ovoviviparity being an adaptation to combat low water temperatures could be rejected for C. bifrons from Windermere.     

The influence of the rate of temperature change on the activation of dormant seeds of Rumex obtusifolius L.

1. One temperature shift from 20 to 30°C in darkness induces 30–40% germination in Rumex obtusifolius seeds. The same germination percentages are found with heat treatment varying between 1 and 6h duration, indicating that the total heat sum of the temperature shift is not important.
2. Germination is greatly enhanced by three consecutive heat shifts of 1h at 30°C separated by 1h periods at 20°C.
3. The seeds are activated to a small extent after a slow warming (+2°Ch^–1) from 20 to 30°C, followed by incubation for 1h at 30°C. Germination is much higher after rapid heating (+10°Ch^–1) to 30°C, followed by 1h incubation at this temperature. Repeated fast heating treatments on four consecutive days enhances germination. Moderately rapid heatings (+3·3°Ch^–1) give intermediate results.
4. The rate of cooling does not influence the germination percentage. Cooling alone cannot induce germination.
5. Heating alone from 15 to 25°C without cooling also activates germination. In this temperature range the seeds are more activated by rapid warming than by slow warming.
6. The ecological relevance of the response to different warming rate is discussed. The insensitivity of seeds to a slow warming might keep deeply buried seeds in a dormant stage.     

Effect of pre- and post-heat shock temperature on the persistence of thermotolerance and heat shock-induced proteins in Listeria monocytogenes

The effect of incubation temperature, before and after a heat shock, on thermotolerance of Listeria monocytogenes at 58°C was investigated. Exposing cells grown at 10°C and 30°C to a heat shock resulted in similar rises in thermotolerance while the increase was significantly higher when cells were grown at 4°C prior to the heat shock. Cells held at 4°C and 10°C after heat shock maintained heat shock-induced thermotolerance for longer than cells held at 30°C. The growth temperature prior to inactivation had negligible effect on the persistence of heat shock-induced thermotolerance. Concurrent with measurements of thermotolerance were measurements of the levels of heat shock-induced proteins. Major proteins showing increased synthesis upon the heat shock had approximate molecular weights of 84, 74, 63, 25 and 19 kDa. There was little correlation between the loss of thermotolerance after the heat shock and the levels of these proteins. Thermotolerance of heat shocked and non-heat shocked cells was described by traditional log-linear kinetics and a model describing a sigmoidal death curve (logistic model). Employing log-linear kinetics resulted in a poor fit to a major part of the data whereas a good fit was achieved by the use of a logistic model.