Improved Temperature-Gradient Incubator and the Maximal Growth Temperature and Heat Resistance of Salmonella

An improved all-metal temperature-gradient incubator produces its gradient by means of a bar permanently installed in a near-vertical position with its lower end in a cool constant-temperature water bath and with thermostatically controlled heaters near its top. Bolts hold the incubator in contact with the temperature-gradient bar, and polyurethane foam insulates the entire assemblage during use. Maximal growth temperatures of 34 representative strains of Salmonella were found to be between 43.2 and 46.2 C. In an agar medium with an initial level of 10⁶ cells per milliliter, no strain survived 50 C for 48 hr. S. senftenberg 775W showed no greater heat resistance at or near 48 C than did other species or other S. senftenberg strains. However, it was considerably more resistant than other strains at 55 C.     

Behavioural thermoregulation and activity patterns in the green sunfish, Lepomis cyanellus.

Behavioural thermoregulation and locomotory activity of green sunfish were examined in a temporal temperature-gradient apparatus. Green sunfish actively avoided temperatures exceeding 30.3 degrees C and below 26.5 degrees C and had a median preferred temperature of 28.2 degrees C. Temperature preference did not vary significantly during the diel period even though locomotory activity patterns were markedly crepuscular. Activity was stimulated by the change in illumination levels at dawn and dusk.     

Temperature-gradient gel electrophoresis. Thermodynamic analysis of nucleic acids and proteins in purified form and in cellular extracts

A temperature-gradient gel electrophoresis technique and its application to the study of structural transitions of nucleic acids and protein-nucleic acid complexes are described. The temperature gradient is established in a slab gel by means of a simple ancillary device for a commercial horizontal gel apparatus. The gradient may be freely selected between 10 and 80 degrees C, and is highly reproducible and linear. In a normal application the biopolymers migrate perpendicular to the temperature gradient so that every individual molecule is at constant temperature throughout electrophoresis. The structural transition of a biopolymer is seen as a continuous band which is retarded or speeded up in the temperature range of the transition. Dissociation processes are mostly irreversible under the conditions of electrophoresis and, therefore, show up as discontinuous transitions from a slow-moving to fast-moving band. As examples the conformational transitions of viroids, double-stranded RNA from reovirus, double-stranded satellite RNA from cucumber mosaic virus and repressor-operator complexes have been studied. It could be shown that by this method dsRNA molecules may be differentiated which differ only in one base-pair, or proteins differing in one amino acid only. As a particular advantage, temperature-gradient gel electrophoresis allows the study of conformational transitions of biopolymers which have not been purified. The biopolymer may either be identified by silver staining as a specific band among many others or, if the study is carried out on nucleic acids, these may be recorded by hybridization with a radioactive probe.     

Alternating Temperatures and Rate of Seed Germination in Lentil

The effect of alternating temperatures on the times taken byseeds of lentil (Lens culinaris Medikus) to germinate was investigatedusing a two-way temperature-gradient plate. Between 5 and 25°C,warmer temperatures increased the rate of germination. Variationamong the individual seeds in the times required for germinationat different constant temperatures within this range were describedwell by a log-normal distribution of thermal times, accumulatedabove a base temperature of 1·5°C. Even with amplitudesas great as 20°C, no effect of alternation per se on thethermal time required for germination was detected—whetherthe cool temperature was applied for 8 or 16 h d^-1. Similarly,in alternating temperature regimes where the minimum temperatureof the diurnal cycle was between 0°C and the base temperature,the thermal times required for germination (where no thermaltime accrued during the periods when temperature was below T_b)were in close agreement with those values provided by the modeldetermined at warmer constant temperatures. However, where theminimum temperature applied was < 0°C the germinationof all but the earliest germinators was delayed beyond modelpredictions, and more so where the sub-zero minimum temperaturewas applied for 16 rather than 8 h d^-1. The results, therefore,contradict the view that alternation in temperature per se reducesthe thermal time required for seed germination. Rather, rateof germination responds instantaneously to current temperature,but prolonged exposure to sub-zero temperatures can result indamage sufficient to delay germination when seeds are returnedto regimes warmer than the base temperature.Copyright 1994,1999 Academic Press Lens culinaris Medikus, lentil, seed germination, alternating temperatures, thermal time, temperature-gradient plate     

Molecular basis for low temperature compartment formation by transitional endoplasmic reticulum of rat liver

The molecular basis for temperature compartment formation was investigated using a cell-free system from rat liver. The donor was from liver slices prelabeled with [³H]acetate. Unlabeled Golgi apparatus membranes were immobilized on nitrocellulose as the acceptor. When transfer was determined as a function of temperature, a transition in transfer activity was observed at low temperatures (≤ 20°C) similar to that seen in vivo. The decrease in transfer efficiency correlated with a decrease in phosphatidylethanolamine and phosphatidylserine content of the transition vesicles formed. By adding lipid mixtures enriched in these lipids to the vesicles, their ability to fuse with the cis Golgi apparatus was reconstituted. These findings provide evidence for a role for lipids in low temperature compartment formation.     

The effect of temperature on growth and reproduction of Scytosiphon complanatus (Fucophyceae) from Greenland

Scytosiphon complanatus from Greenland was grown under long-day conditions on a temperature-gradient device with a temperature range from 5.4°C to 31.8°C. Growth was optimal between 16.0°C and 20.9°C. In a four week experimental period at 5.4°C and 7.5°C growth was slow and not measurable. The inoculated germlings died at temperatures between 24.0°C and 27.5°C. Under all temperatures the prostrate systems, knot filaments or ralfsioid thalli, as well as the parenchymatous macrothalli remained sterile during the experimental period. Prolongation of the growth period showed that formation of swarmers was prevented at temperatures above 18.6°C. The geographic distribution is discussed in relation to these results.     

Low temperature compartment formation in feline immunodeficiency virus-infected and uninfected feline kidney cells

Summary This study was to determine if feline immunodeficiency virus (FIV)-infected and uninfected Crandall feline kidney (CRFK) cells exhibited a low temperature (16°C) block in membrane trafficking between transitional endoplasmic reticulum and Golgi apparatus represented by intermediate compartment formation. Cells were cultured at different temperatures and membrane changes involving the Golgi apparatus and Golgi apparatus-associated membrane structures were monitored by electron microscopy and quantitated. With 30 min of incubation, membranes of the Golgi apparatus stack increased in amount at temperatures of 16°C and below compared to temperatures above 18°C. The increase was greatest along the major polarity axis as evidenced by an increased stack height. Neither the number of cisternae per stack nor the average stack diameter (width) was affected by temperature. The response was maximal between 15 and 30 min of low temperature treatment of the cells. Results with cells infected and uninfected with feline immunodeficiency virus were similar. The increase in stack height was due primarily to an increase of membranes at the cis face (cis Golgi apparatus network). At 18°C, membranes of the trans Golgi apparatus network accumulated suggesting that import from the cis Golgi network could proceed at this temperature, whereas exit from the trans Golgi network was still at least partially blocked. Also increased at 16°C and below were numbers of transition vesicles in the space between the Golgi apparatus and the transitional endoplasmic reticulum associated with the cis Golgi apparatus face. The results suggested interruption of the orderly flux of membranes into the Golgi apparatus at 16°C and below. Moreover, the block appeared to be reversible. Upon transfer from 16°C to 37°C, there was a time-dependent decrease in the accumulations of cis compartment membrane accompanied by a corresponding equivalent increase in the membranes of the trans Golgi apparatus compartment.     

Thermal preference of Arctic charr, <Emphasis Type="Italic">Salvelinus alpinus</Emphasis>, and brown trout, <Emphasis Type="Italic">Salmo trutta</Emphasis> – implications for their niche segregation

This study tested a new experimental apparatus to estimate thermal preferences of fish. The apparatus was designed to minimise the effect of the thermal history of the fish and allow for easy feeding. The set-up consisted of two connected sections of an aquarium, both receiving an excess of food, with slightly different water temperatures. Initially, the fish spent most of its time in one of the sections, but when the temperatures were slowly increased (or decreased), the fish spent increasingly more time in the other. The temperature at which the fish spent equal time in both sections was defined as the preferred temperature. Brown trout, Salmo trutta, preferred the reported optimal temperature for growth of the species. However, Arctic charr, Salvelinus alpinus, selected a significantly lower temperature than its optimal temperature for growth and thus contradicted the general view of good correlation between the optimal temperature for growth and preferred temperature of fish. The reason for this may be that charr is optimising its growth efficiency instead of its growth rate. Individuals that utilise a limited resource in an optimal way, by selecting a temperature where the growth efficiency is maximised, will possibly be favoured. Several factors affect the distribution of fish in lakes, but the difference in thermal preference between charr and trout might partly explain the frequently observed niche segregation of these two species in Scandinavian lakes.     

Interactive effects of rutin and constant versus alternating temperatures on performance ofManduca sexta caterpillars

The effects of different concentrations of rutin and constant temperature (20 °C) versus alternating temperatures (23∶15 °C) on growth, molting and food utilization efficiencies of third instar tobacco hornworms (Manduca sexta) were determined. Relative consumption rate (RCR) and relative growth rate (RGR) were significantly higher for larvae at the alternating thermal regime compared to those at the constant (representing the average) temperature. With increasing concentrations of rutin, the negative effect of rutin on RCR and RGR increased for the larvae in the alternating thermal regime; however, at the constant temperature, rutin had little effect. The alternating thermal regime promoted synchrony in the timing of spiracle apolysis (the earliest morphological marker of molt). Rutin disrupted that synchrony. I discuss how patterns of host plant resistance may be altered with a decrease, in amplitude of diurnal temperatures (as has been documented recently for temperate regions) through the uncoupling of herbivore performance and allelochemical concentration. I conclude that simultaneous consideration of fluctuating temperatures and allelochemicals is advisable when assessing the effects of temperature and allelochemicals on performance of insect herbivores because interactive effects between temperature and dietary components occur and perhaps are common.     

Growth of Rhizobium japonicum Strains at Temperatures Above 27 degrees C

A study was conducted to examine the growth responses of different Rhizobium japonicum strains to increasing temperatures, determine the degree of variability among strains in those responses, and identify temperature-related growth characteristics that could be used to select temperature-tolerant strains. Each of 42 strains was grown in liquid culture for 96 h at 19 incubation temperatures ranging from 27.4 to 54.1 degrees C in a temperature gradient apparatus. Growth was estimated by measuring the change in optical density over time. Strains differed in their responses to increasing temperatures. Three characteristic temperatures were determined for each strain: the temperature giving the maximum optical density at 96 h (optimum temperature), the maximum temperature allowing a continuous increase in optical density during the 96-h period (maximum permissive temperature), and the maximum temperature allowing growth of the cultures after they were transferred to a uniform incubation temperature of 28 degrees C (maximum survival temperature). The three characteristic temperatures varied among strains and had the following ranges: optimum temperature, from 27.4 to 35.2 degrees C; maximum permissive temperature, from 29.8 to 38.0 degrees C; and maximum survival temperature, from 33.7 to 48.7 degrees C. Significant positive correlations were found between maximum permissive temperature and optimum temperature and between maximum permissive temperature and maximum survival temperature. Eight strains which had the highest maximum permissive temperature, optimum temperature, and maximum survival temperature were considered tolerant of high temperatures and were able to grow at temperatures higher than those previously reported for the most tolerant R. japonicum strains. The strains were of diverse geographical origin, but the response to high temperatures was not related to their origin. Evaluation of the temperature responses in pure culture may be useful in the search for R. japonicum strains better suited to environments in which high soil temperature is a limiting factor.     

The response to temperature gradients of scorpions from mesic and xeric habitats

• 1.1. The thermal preference of four scorpion species—two from xeric habitats and two from mesic habitats—was observed in a temperature-gradient.
• 2.2. The two scorpions from mesic habitats responded positively to lower temperatures. The xeric species stayed longer in higher temperatures.
• 3.3. In each pair of scorpion species studied here (the mesic and the xeric species), one of them responded to a more limited temperature range whereas the other to a wider range.
• 4.4. The significance of these findings is discussed in relation to the microclimate in the scorpion's microhabitat and to their dispersal pattern.

     

Identification of an intermediate compartment involved in protein transport from endoplasmic reticulum to Golgi apparatus

We have studied the role of a previously described tubulovesicular compartment near the cis-Golgi apparatus in endoplasmic reticulum (ER)-to-Golgi protein transport by light and immunoelectron microscopy in Vero cells. The compartment is defined by a 53-kDa transmembrane protein designated p53. When transport of the vesicular stomatitis virus strain ts045 G protein was arrested at 39.5 degrees C, the G protein accumulated in the ER but had access to the p53 compartment. At 15 degrees C, the G protein was exported from the ER into the p53 compartment which formed a compact structure composed of vesicular and tubular profiles in close proximity to the Golgi. Upon raising the temperature to 32 degrees C, the G protein migrated through the Golgi apparatus while the p53 compartment resumed its normal structure again. These results establish the p53 compartment as the 15 degrees C intermediate of the ER-to-Golgi protein transport pathway.     

Physiological adaptation ofMytilus edulis to cyclic temperatures

Summary Mytilus edulis adapted to cyclic temperatures by reducing the amplitude of response of oxygen consumption and filtration rate over a period of approximately two weeks, and thereby increasing their independence of temperature within the range of the fluctuating regime. When acclimated to cyclic temperature regimes within the range from 6 to 20°C, the metabolic and feeding rates, measured at different temperatures in the cycle, were not significantly different from the adapted response to equivalent constant temperatures.Physiological adaptation ofMytilus edulis to different thermal environments was reflected in their metabolic and feeding rate-temperature curves. Animals subjected to marked diel fluctuations in environmental temperature showed an appropriate region of temperature-independence, whereas animals from a population not experiencing large diel temperature fluctuations showed no region of temperature-independence.In a fluctuating thermal environment which extended above the normal environmental maxima, respiratory adaptation occurred at higher temperatures than was possible in a constant thermal environment. The feeding rate was also maintained at higher temperatures in a cyclic regime than was possible under constant thermal conditions. This represented a shortterm extension of the zone of activity in a fluctuating thermal environment. The net result of these physiological responses to high cyclic and constant temperatures has been assessed in terms of scope for growth. Animals acclimated to cyclic temperatures between 21 and 29°C had a higher scope for growth at 29°C and were less severely stressed than those maintained at the constant temperature of 29°C.     

Modeling Surface Growth of Escherichia coli on Agar Plates

Surface growth of Escherichia coli cells on a membrane filter placed on a nutrient agar plate under various conditions was studied with a mathematical model. The surface growth of bacterial cells showed a sigmoidal curve with time on a semilogarithmic plot. To describe it, a new logistic model that we presented earlier (H.Fujikawa et al., Food Microbiol. 21:501-509, 2004) was modified. Growth curves at various constant temperatures (10 to 34°C) were successfully described with the modified model (model III). Model III gave better predictions of the rate constant of growth and the lag period than a modified Gompertz model and the Baranyi model. Using the parameter values of model III at the constant temperatures, surface growth at various temperatures was successfully predicted. Surface growth curves at various initial cell numbers were also sigmoidal and converged to the same maximum cell numbers at the stationary phase. Surface growth curves at various nutrient levels were also sigmoidal. The maximum cell number and the rate of growth were lower as the nutrient level decreased. The surface growth curve was the same as that in a liquid, except for the large curvature at the deceleration period. These curves were also well described with model III. The pattern of increase in the ATP content of cells grown on a surface was sigmoidal, similar to that for cell growth. We discovered several characteristics of the surface growth of bacterial cells under various growth conditions and examined the applicability of our model to describe these growth curves.     

Identification of the 16 degrees C compartment of the endoplasmic reticulum in rat liver and cultured hamster kidney cells

In many systems transfer between the endoplasmic reticulum and the Golgi apparatus is blocked at temperatures below 16 degrees C. In virus-infected cells in culture, a special membrane compartment is seen to accumulate. Our studies with rat liver show a similar response to temperature both in situ with slices and in vitro with isolated transitional endoplasmic reticulum fractions. With isolated transitional endoplasmic reticulum fractions, when incubated in the presence of nucleoside triphosphate and a cytosol fraction, temperature dependent formation of vesicles occurred with a Q10 of approximately 2 but was apparent only at temperatures greater than 12 degrees C. A similar response was seen in situ at 12 degrees C and 16 degrees C where fusion of transition vesicles with cis Golgi apparatus, but not their formation, was blocked and transition vesicles accumulated in large numbers. At 18 degrees C and below and especially at 8 degrees C and 12 degrees C, the cells responded by accumulating smooth tubular transitional membranes near the cis Golgi apparatus face. With cells and tissue slices at 20 degrees C neither transition vesicles nor the smooth tubular elements accumulated. Those transition vesicles which formed at 37 degrees C were of a greater diameter than those formed at 4 degrees C both in situ and in vitro. The findings show parallel responses between the temperature dependency of transition vesicle formation in vitro and in situ and suggest that a subpopulation of the transitional endoplasmic reticulum may be morphologically and functionally homologous to the 16 degrees C compartment observed in virally-infected cell lines grown at low temperatures.     

A Compartment Model of the Effect of Early-season Temperatures on Potential Size and Growth of 'Delicious' Apple Fruits

A compartmental growth model was developed to describe expansionof ‘Delicious’ apple fruit diameter and the effectof early-season temperatures on potential size at harvest. Themodel was based on the assumption that growth may be describedas a function of transfer between two conceptual compartments.Under this scheme, the first compartment represented all tissuecontributing to the setting of potential fruit size (determinedas the integral of its output) whereas the second compartmentrepresented all other fruit tissue whose growth actualized thatpotential. Expansion of both compartments was assumed to havea temperature response with an optimum, whereas an aging processwith an asymptotic temperature response controlled transferto the second compartment. Model parameters were estimated byfitting to data from controlled environment experiments in whichearly-season temperature conditions were varied. Predicted fruitgrowth curves showed close agreement with measured diameterdata. The results were consistent with a two-fold impact ofearly-season temperatures on apple fruit size: an immediate,direct effect on growth rate and an enduring effect, mediatedthrough fruit cell number or resource allocation to young fruit,reflecting the establishment of a potential that subsequentgrowth actualizes.Copyright 1999 Annals of Botany Company. Malus domesticaBorkh., apples, ‘Delicious’, fruit growth, models, temperature, potential size, cell division.     

Membrane flow in plants: preparation and kinetics of labelling of plasma membranes from growing pollen tubes of tobacco

Summary Growing pollen tubes of tobacco germinated in suspension culture, were labelled with [³H]leucine and after varying times of chase with unlabelled leucine at 23, 16, or 4°C, were separated into plasma membrane-enriched and plasma membrane-depleted fractions by aqueous two-phase partition. At 23°C, the specific radioactivity of the plasma membrane increased with time to a maximum at 60 min. At 16°C and 4°C, labelling of the plasma membrane was respectively 40% and 10% that at 23°C. However, if labelling was at 23°C and subsequent transfer was at 4°C, plasma membrane labelling was much less affected and labelling of the plasma membrane was 60% that at 23°C. Additionally, quantitation of various morphological parameters revealed no accumulations of 50–70 nm transition vesicles in the space between endoplasmic reticulum and cis Golgi apparatus that might suggest formation of a low temperature compartment similar to those described for mammalian cells and tissues. Similarly, growth of pollen tubes was reduced but not blocked even at temperatures of 12°C. The results suggest that tube elongation is accompanied by a steady state flow of membranes to the cell surface that is relatively insensitive to interruption by low temperatures. Whereas leucine incorporation is reduced by low temperature even at 16°C, the flow pathway to the cell surface, including the endoplasmic reticulum to Golgi apparatus transfer step, as well as elongation growth does not exhibit a pronounced low temperature block in this tip growing system.     

The effect of daily exposure to low hardening temperature on plant vital activity

Phenomenological responses of plants to daily short-term exposure to low hardening temperature was studied under chamber and field conditions. Experiments were carried out on cucumber (Cucumis sativus L.), barley (Hordeum vulgare L.), marigolds (Tagetes L.), and petunia (Petunia × hybrida) plants. The obtained data demonstrated a similar pattern of response in all studied plant species to different variants of exposure to low hardening temperature. The main features of plant response to daily short-term exposure to low hardening temperature include: a higher increment in cold tolerance (cf. two-or threefold increase relative to constant low hardening temperature) that peaked on day 5 (cf. day 2 at constant low hardening temperature) and was maintained for 2 weeks (cf. 3–4 days at constant low hardening temperature); a simultaneous increase in heat tolerance (cf. twofold relative to constant low hardening temperature) maintained over a long period (cf. only in the beginning of the exposure to constant low hardening temperature); a sharp drop in the subsequent cold tolerance after plant incubation in the dark (cf. a very low decrease in cold tolerance following the exposure to constant low hardening temperature); a combination of high cold tolerance and high photochemical activity of the photosynthetic apparatus (cf. a low non-photochemical quenching at constant low hardening temperature); and the capacity to increase cold tolerance in response to repeated short-term exposures to low hardening temperature in plants grown outdoors (cf. a gradual increase after repeated exposure to constant low hardening temperature). Possible mechanisms underlying the plant response to daily short-term exposure to low temperature are proposed.     

Molecular cloning, characterization, subcellular localization and dynamics of p23, the mammalian KDEL receptor

We have isolated a cDNA clone (mERD2) for the mammalian (bovine) homologue of the yeast ERD2 gene, which codes for the yeast HDEL receptor. The deduced amino acid sequence bears extensive homology to its yeast counterpart and is almost identical to a previously described human sequence. The sequence predicts a very hydrophobic protein with multiple membrane spanning domains, as confirmed by analysis of the in vitro translation product. The protein encoded by mERD2 (p23) has widespread occurrence, being present in all the cell types examined. p23 was localized to the cis-side of the Golgi apparatus and to a spotty intermediate compartment which mediates ER to Golgi transport. A majority of the intracellular staining could be accumulated in the intermediate compartment by a low temperature (15 degrees C) or brefeldin A. During recovery from these treatments, the spotty intermediate compartment staining of p23 was shifted to the perinuclear staining of the Golgi apparatus and tubular structures marked by p23 were observed. These tubular structures may serve to mediate transport between the intermediate compartment and the Golgi apparatus.     

Double-stranded cucumovirus associated RNA 5: experimental analysis of necrogenic and non-necrogenic variants by temperature-gradient gel electrophoresis.

Cucumber mosaic virus (CMV) and peanut stunt virus (PSV) each contain a fifth major RNA in the size range of 334 to 393 nucleotides. This fifth RNA is a satellite capable of modulating the expression of viral disease symptoms. It is present in infected tissue in single-stranded and double-stranded form. Nucleotide sequence variants of the double-stranded CMV-associated RNA 5 (dsCARNA 5) and PSV-associated RNA 5 (dsPARNA 5) were analysed by temperature-gradient gel electrophoresis. Gels were 5% polyacrylamide, containing 8 M urea in 8.9 mM Tris-borate buffer, with temperature differences of 25-40 degrees C establishing gradients either perpendicular or parallel to the direction of the electric field. For dsCARNA 5 two characteristic transitions were detected with increasing temperature: at temperatures between 40 degrees C and 46 degrees C a drastic retardation in electrophoretic mobility induced by partial dissociation of the duplex structure from the ends and at temperatures above 52 degrees C an abrupt increase in mobility due to complete strand dissociation. dsPARNA 5 exhibited both transitions at up to 10 degrees C higher temperatures and an additional retardation between the transitions mentioned. Seven different variants of dsCARNA 5, 4 necrogenic and 3 non-necrogenic, were analysed. Some showed only one single band, others gave rise to up to six well separated bands corresponding to six molecular species. From all experimental results a correlation between the temperature of the retardation transition and the necrogenicity of CARNA 5 was derived. The diagnostic application of the temperature-gradient gel analysis in agriculture, particularly for the use of non-necrogenic variants as biological control agents to impede CMV-infections, is discussed.