EVOLUTIONARY ADAPTATION TO TEMPERATURE II. THERMAL NICHES OF EXPERIMENTAL LINES OF ESCHERICHIA COLI

Groups of replicated lines of the bacterium Escherichia coli were propagated for 2,000 generations at constant 32, 37, or 42°C, or in an environment that alternated between 32 and 42°C. Here, we examine the performance of each group across a temperature range of 12-44°C measuring the temperatures over which each line can maintain itself in serial dilution culture (the thermal niche). Thermal niche was not affected by selection history: average lower and upper limits remained about 19 and 42°C for all groups. In addition, no significant differences among groups were observed in rate of extinction at more extreme temperatures. Within the thermal niche, we measured the mean fitness of the evolved groups relative to their common ancestor. Increases in mean fitness were temperature specific, with the largest increase for each group occurring near its selected temperature. Thus, the temperature at which mean fitness relative to the ancestor was greatest (the thermal optimum) diverged by about 10°C for the groups selected at constant 32°C versus constant 42°C. Tradeoffs in relative fitness (decrements relative to the ancestor elsewhere within the thermal niche) did not necessarily accompany fitness improvements, although tradeoffs were observed for a few of the lines. We conclude that adaptation in this system was quite temperature specific, but substantial divergence among groups in thermal optima had little effect on the limits of their thermal niches and did not necessarily involve tradeoffs in fitness at other temperatures.     

EVOLUTIONARY ADAPTATION TO TEMPERATURE. IV. ADAPTATION OF ESCHERICHIA COLI AT A NICHE BOUNDARY

Following an environmental change, the course of a population's adaptive evolution may be influenced by environmental factors, such as the degree of marginality of the new environment relative to the organism's potential range, and by genetic factors, including constraints that may have arisen during its past history. Experimental populations of bacteria were used to address these issues in the context of evolutionary adaptation to the thermal environment. Six replicate lines of Escherichia coli (20°C group), founded from a common ancestor, were propagated for 2000 generations at 20°C, a novel temperature that is very near the lower thermal limit at which it can maintain a stable population size in a daily serial transfer (100-fold dilution) regime. Four additional groups (32/20, 37/20, 42/20, and 32–42/20°C groups) of six lines, each with 2000 generation selection histories at different temperatures (32, 37, 42, and daily alternation of 32 and 42°C), were moved to the same 20°C environment and propagated in parallel to ascertain whether selection histories influence the adaptive response in this novel environment. Adaptation was measured by improvement in fitness relative to the common ancestor in direct competition experiments conducted at 20°C. All five groups showed improvement in relative fitness in this environment; the mean fitness of the 20°C group after 2000 generations increased by about 8%. Selection history had no discernible effect on the rate or final magnitude of the fitness responses of the four groups with different histories after 2000 generations. The correlated fitness responses of the 20°C group were measured across the entire thermal niche. There were significant tradeoffs in fitness at higher temperatures; for example, at 40°C the average fitness of the 20°C group was reduced by almost 20% relative to the common ancestor. We also observed a downward shift of 1–2°C in both the upper and lower thermal niche limits for the 20°C selected group. These observations are contrasted with previous observations of a markedly greater rate of adaptation to growth near the upper thermal limit (42°C) and a lack of trade-off in fitness at lower temperatures for lines adapted to that high temperature. The evolutionary implications of this asymmetry are discussed.     

EVOLUTIONARY ADAPTATION TO TEMPERATURE. IX. PREADAPTATION TO NOVEL STRESSFUL ENVIRONMENTS OF ESCHERICHIA COLI ADAPTED TO HIGH TEMPERATURE

Abstract Stressful environments may be considered as those that reduce fitness, sometimes due in part to the increased metabolic expenditure required to sustain life. Direct adaptation to a stressor is expected to increase fitness and reduce maintenance metabolism, with the latter leading to increased biomass production. In this study, we test the general hypothesis that such adaptation to one stressor can preadapt organisms to novel stressful environments. Six lines of Escherichia coli propagated for 2000 generations at 41–42°C (42 group), a stressful temperature, were compared to six control lines propagated for 2000 generations at 37°C (37 group) and to the common ancestor of both groups. We assayed biovolume yield (a measure of growth efficiency) and competitive fitness in the 42 group's selective high temperature environment as well as five novel stressful environments–acid, alkali, ethanol, high osmolarity and peroxide. As previously reported, at high temperature the 42 group had both higher yield and fitness than the 37 group and ancestor. In the novel environments, the 42 group generally produced yields higher than the 37 group (and marginally higher than the ancestor), but we found no differences in competitive fitness among the 37 and 42 groups and the ancestor. We also found that the performance of lines within groups was not correlated across stressful environments for either yield or relative fitness. Because previous adaptation to one stressor did not improve our measure of Darwinian fitness in novel stressful environments, we conclude that the 42 group shows no useful preadaptation, or cross‐tolerance, to these types of environments.     

EVOLUTIONARY ADAPTATION TO TEMPERATURE. III. ADAPTATION OF ESCHERICHIA COLI TO A TEMPORALLY VARYING ENVIRONMENT

Six lines of the bacterium Escherichia coli were propagated for 2,000 generations in a temporally varying environment. The imposed environmental regime consisted of alternating days at 32°C and 42°C, with rapid transitions between them. These derived lines are competitively superior to their ancestor in this variable temperature regime. We also measured changes in the fitness of these lines, relative to their common ancestor, in both the constant (32°C and 42°C) and transition (from 32°C to 42°C and from 42°C to 32°C) components of this temporally varying environment, to determine whether the bacteria had adapted to the particular constant temperatures or the transitions between them, or both. The experimentally evolved lines had significantly improved fitness in each of the constant environmental components (32°C and 42°C). However, the experimental lines had not improved in making the sudden temperature transitions that were a potentially important aspect of the temporally variable environment. In fact, fitness in making at least one of the transitions (between 32°C and 42°C) unexpectedly decreased. This reduced adaptation to the abrupt transitions between these temperatures is probably a pleiotropic effect of mutations that were responsible for the increased fitness at the component temperatures. Among the six experimental lines, significant heterogeneity occurred in their adaptation to the constant and transition components of the variable environment.     

EVOLUTIONARY ADAPTATION TO TEMPERATURE. V. ADAPTIVE MECHANISMS AND CORRELATED RESPONSES IN EXPERIMENTAL LINES OF ESCHERICHIA COLI

We previously demonstrated temperature-specific genetic adaptation in experimental lines of Escherichia coli. Six initially identical populations were propagated for 2000 generations under each of five regimes: constant 20°C, 32°C, 37°C, and 42°C, and a daily switch between 32°C and 42°C. Glucose was the sole carbon source in all cases. Here, we examine the physiological bases of adaptation to determine whether the same mechanisms evolved among the replicate lines within each thermal regime and across different regimes. Specifically, we investigate whether changes in glucose transport may account for the temperature-specific adaptation. We compared each line's direct response of fitness to glucose with its correlated response to maltose; glucose and maltose enter the cell by different pathways, but their catabolism is identical. Except for lines maintained at the ancestral temperature (37°C), almost all derived lines had significantly different fitnesses (relative to their common ancestor) in glucose and maltose, supporting the hypothesis that adaptation involved changes in glucose transport. An alternative explanation, that maltose transport decayed by genetic drift, appears unlikely for reasons that are discussed. Although most lines showed evidence of temperature-specific adaptation in glucose transport, several different mechanisms may underlie these improvements, as indicated by heterogeneity in correlated responses (across temperatures and substrates) among replicate lines adapted to the same regime. This heterogeneity provides a latent pool of genetic variation for responding to environmental change.     

EVOLUTIONARY ADAPTATION TO TEMPERATURE. VI. PHENOTYPIC ACCLIMATION AND ITS EVOLUTION IN ESCHERICHIA COLI

Acclimation refers to reversible, nongenetic changes in phenotype that are induced by specific environmental conditions. Acclimation is generally assumed to improve function in the environment that induces it (the beneficial acclimation hypothesis). In this study, we experimentally tested this assumption by measuring relative fitness of the bacterium Escherichia coli acclimated to different thermal environments. The beneficial acclimation hypothesis predicts that bacteria acclimated to the temperature of competition should have greater fitness than do bacteria acclimated to any other temperature. The benefit predicted by the hypothesis was found in only seven of 12 comparisons; in the other comparisons, either no statistically demonstrable benefit was observed or a detrimental effect of acclimation was demonstrated. For example, in a lineage evolutionarily adapted to 37°C, bacteria acclimated to 37°C have a higher fitness at 32°C than do bacteria acclimated to 32°C, a result exactly contrary to prediction; acclimation to 27°C or 40°C prior to competition at those temperatures confers no benefit over 37°C acclimated forms. Consequently, the beneficial acclimation hypothesis must be rejected as a general prediction of the inevitable result of phenotypic adjustments associated with new environments. However, the hypothesis is supported in many instances when the acclimation and competition temperatures coincide with the historical temperature at which the bacterial populations have evolved. For example, when the evolutionary temperature of the population was 37°C, bacteria acclimated to 37°C had superior fitness at 37°C to those acclimated to 32°C; similarly, bacteria evolutionarily adapted to 32°C had a higher fitness during competition at 32°C than they did when acclimated to 37°C. The more surprising results are that when the bacteria are acclimated to their historical evolutionary temperature, they are frequently competitively superior even at other temperatures. For example, bacteria that have evolved at either 20°C or 32°C and are acclimated to their respective evolutionary temperatures have a greater fitness at 37°C than when they are acclimated to 37°C. Thus, acclimation to evolutionary temperature may, as a correlated consequence, enhance performance not only in the evolutionary environment, but also in a variety of other thermal environments.     

EVOLUTIONARY ADAPTATION TO TEMPERATURE. VII. EXTENSION OF THE UPPER THERMAL LIMIT OF ESCHERICHIA COLI

What factors influence the ability of populations to adapt to extreme environments that lie outside their current tolerance limits? We investigated this question by exposing experimental populations of the bacterium Escherichia coli to lethally high temperatures. We asked: (1) whether we could obtain thermotolerant mutants with an extended upper thermal limit by this selective screen; (2) whether the propensity to obtain thermotolerant mutants depended on the prior selective history of the progenitor genotypes; and (3) how the fitness properties of these mutants compared to those of their progenitors within the ancestral thermal niche. Specifically, we subjected 15 independent populations founded from each of six progenitors to 44°C; all of the progenitors had upper thermal limits between about 40°C and 42°C. Two of the progenitors were from populations that had previously adapted to 32°C, two were from populations adapted to 37°C, and two were from populations adapted to 41–42°C. All 90 populations were screened for mutants that could survive and grow at 44°C. We obtained three thermotolerant mutants, all derived from progenitors previously adapted to 41–42°C. In an earlier study, we serendipitously found one other thermotolerant mutant derived from a population that had previously adapted to 32°C. Thus, prior selection at an elevated but nonlethal temperature may predispose organisms to evolve more extreme thermotolerance, but this is not an absolute requirement. It is evidently possible to obtain mutants that tolerate more extreme temperatures, so why did they not become prevalent during prior selection at 41–42°C, near the upper limit of the thermal niche? To address this question, we measured the fitness of the thermotolerant mutants at high temperatures just within the ancestral niche. None of the four thermotolerant mutants had an advantage relative to their progenitor even very near the upper limit of the thermal niche; in fact, all of the mutants showed a noticeable loss of fitness around 41°C. Thus, the genetic adaptations that improve competitive fitness at high but nonlethal temperatures are distinct from those that permit tolerance of otherwise lethal temperatures.     

Indirect selection of thermal tolerance during experimental evolution of Drosophila melanogaster

Natural selection alters the distribution of a trait in a population and indirectly alters the distribution of genetically correlated traits. Long‐standing models of thermal adaptation assume that trade‐offs exist between fitness at different temperatures; however, experimental evolution often fails to reveal such trade‐offs. Here, we show that adaptation to benign temperatures in experimental populations of Drosophila melanogaster resulted in correlated responses at the boundaries of the thermal niche. Specifically, adaptation to fluctuating temperatures (16–25°C) decreased tolerance of extreme heat. Surprisingly, flies adapted to a constant temperature of 25°C had greater cold tolerance than did flies adapted to other thermal conditions, including a constant temperature of 16°C. As our populations were never exposed to extreme temperatures during selection, divergence of thermal tolerance likely reflects indirect selection of standing genetic variation via linkage or pleiotropy. We found no relationship between heat and cold tolerances in these populations. Our results show that the thermal niche evolves by direct and indirect selection, in ways that are more complicated than assumed by theoretical models.     

Morphological Changes of <Emphasis Type="Italic">Pseudomonas pseudoalcaligenes</Emphasis> in Response to Temperature Selection

Adaptation to novel environments usually entails morphological changes. The cell morphology of six experimental populations of Pseudomonas pseudoalcaligenes and their common ancestor were examined with scanning electron microscopy (SEM). The six experimental populations were propagated under different temperatures for 10 months: three of them cultured at constant normal temperature (35°C) forming the control group, and the other three cultured at incremental higher temperatures (from 41° to 47°C) as the HT group. SEM showed the deformed and elongated cells in the 6-h cultures of both ancestral and control populations at 45°C, indicating that 45°C is stressful for the ancestral and the control populations. In contrast, the HT populations retained normal cell shape in the 6-h cultures at both 35°C and 45°C. The mean cell volumes of control and HT populations increased 29% and 34%, respectively, relative to the ancestor at their respective thermal regimens, suggestion that the culturing conditions might favor larger cells. Received: 27 March 2002 / Accepted: 30 April 2002     

Temperature fluctuations during development reduce male fitness and may limit adaptive potential in tropical rainforest Drosophila

Understanding the potential for organisms to tolerate thermal stress through physiological or evolutionary responses is crucial given rapid climate change. Although climate models predict increases in both temperature mean and variance, such tolerances are typically assessed under constant conditions. We tested the effects of temperature variability during development on male fitness in the rainforest fly Drosophila birchii, by simulating thermal variation typical of the warm and cool margins of its elevational distribution, and estimated heritabilities and genetic correlations of fitness traits. Reproductive success was reduced for males reared in warm (mean 24 °C) fluctuating (±3 °C) vs. constant conditions but not in cool fluctuating conditions (mean 17 °C), although fluctuations reduced body size at both temperatures. Male reproductive success under warm fluctuating conditions was similar to that at constant 27 °C, indicating that briefly exceeding critical thermal limits has similar fitness costs to continuously stressful conditions. There was substantial heritable variation in all traits. However, reproductive success traits showed no genetic correlation between treatments reflecting temperature variation at elevational extremes, which may constrain evolutionary responses at these ecological margins. Our data suggest that even small increases in temperature variability will threaten tropical ectotherms living close to their upper thermal limits, both through direct effects on fitness and by limiting their adaptive potential.     

Prior evolution in stochastic versus constant temperatures affects RNA virus evolvability at a thermal extreme

It is unclear how historical adaptation versus maladaptation in a prior environment affects population evolvability in a novel habitat. Prior work showed that vesicular stomatitis virus (VSV) populations evolved at constant 37°C improved in cellular infection at both 29°C and 37°C; in contrast, those evolved under random changing temperatures between 29°C and 37°C failed to improve. Here, we tested whether prior evolution affected the rate of adaptation at the thermal‐niche edge: 40°C. After 40 virus generations in the new environment, we observed that populations historically evolved at random temperatures showed greater adaptability. Deep sequencing revealed that most of the newly evolved mutations were de novo. Also, two novel evolved mutations in the VSV glycoprotein and replicase genes tended to co‐occur in the populations previously evolved at constant 37°C, whereas this parallelism was not seen in populations with prior random temperature evolution. These results suggest that prior adaptation under constant versus random temperatures constrained the mutation landscape that could improve fitness in the novel 40°C environment, perhaps owing to differing epistatic effects of new mutations entering genetic architectures that earlier diverged. We concluded that RNA viruses maladapted to their previous environment could “leapfrog” over counterparts of higher fitness, to achieve faster adaptability in a novel environment.     

Thermal inhibition of repair of methylmethane sulfonate-damaged DNA in chick embryo fibroblasts

Chick embryo fibroblasts were treated with the monofunctional alkylating agent methylmethane sulfonate at various concentrations for 1 h at 42°C, rinsed and then incubated post-treatment at various temperatures at which the kinetics of alkali-labile bond disappearance was followed. Growth experiments showed that these cells grew similarly at temperatures of either 37°C or 42°C. Repair as assessed by removal of alkali-labile bond was also similar for postincubation in the temperature range 37–42°C for damage due to methylmethane sulfonate treatment at concentrations less than 1.5 mM. When the postincubation temperature was raised higher than 42.5–43°C, this type of repair was stopped. The normal internal body temperature of adult chickens is about 41.6°C. Hence the present finding indicates that chick cells are much more severely restricted in DNA repair at temperatures above normal than are mammalian cells, which can function in this respect for several deg. C above 37°C.     

Evolution of total net fitness in thermal lines: Drosophila subobscura likes it 'warm'

Fisher's fundamental theorem states that heritable variation for net fitness sets a limit to the rate of response to natural selection. How will temperate (i.e. cold‐tolerant) species cope with contemporary rapid global warming? Using three‐fold replicated lines of Drosophila subobscura that had been allowed to evolve for 4 years (between 32 and 59 generations) at 13 °C (cold), 18 °C (the supposed optimum temperature), and 22 °C (warm) I assess here how net fitness changes according to thermal environments. Net fitness was estimated following the classical approach in population genetics of competing over a number of generation in outbred experimental populations multiple wild‐type O chromosomes (homologous to arm 3R in D. melanogaster) independently derived from each base thermal stock in an otherwise homogeneous genetic background against a balancer chromosome. Warm‐adapted populations (‘warm‐adapted O chromosomes’) performed comparatively well at all tested temperatures. However, net fitness was severely reduced in cold‐adapted populations when transferred to warmer conditions. It seems, therefore, that thermal fitness breath for D. subobscura flies is positively associated to temperature. These findings are discussed in relation to the fast world‐wide clinal shifts in the frequency of genetic markers correlated with current climate change.     

Temperature dependence of capacitation in bat sperm monitored by zona-free hamster ova

The temperature dependence of capacitation in bat sperm (Myotis lucifugus lucifugus) was studied by monitoring fertilizations rates of zona-free hamster ova at different temperatures. Spermatozoa were cultured in BWW medium at temperatures 4°C, 24°C, 32°C, 42°C, and 55°C from 0–24 hr. Activation of sperm could be determined visually due to the change in movement seen through light microscopy. Activation was later confirmed by higher rates of fertilization. Preincubation of the bat sperm was found to have a direct effect on the success of penetration of the zona-free hamster ova. Holding bat spermatozoa at low temperature for long intervals allowed them to remain motile but unable to fertilize. Sperm are not irreversibly damaged, however, and activation, when the temperature is increased to 32°C, is faster than when sperm are intitially put at 32°C, resulting in good fertilization rates.     

Temperature Ranges,Growth Optima,and Growth Rates of Spiroplasma (Spiroplasmataceae,class Mollicutes) Species

A new method was developed for determination of the doubling times of spiroplasmas. In this procedure, the time required for medium acidification of tubes in tenfold dilution series was recorded. Sixty-four spiroplasma strains, representing 24 groups and 11 subgroups, were studied. Eight strains representing putative new groups were also included in the study. Doubling times at 5, 10, 15, 20, 25, 30, 32, 37, 41, and 43°C were determined. The range of temperatures for spiroplasma growth was 5°–41°C. Twenty-three spiroplasmas had optima of 30°C, 29 had optima of 32°C, and 13 had optima of 37°C. The fastest growing spiroplasma was the MQ-4 strain (group XI), with a doubling time at optimal temperature of 0.6 h. The slowest was the Jamaican corn stunt strain B655 (subgroup I-3), with an optimal doubling time of 36.7 h. Spiroplasma strain B31 (group IV) had the widest range (5°–41°C), while the DW-1 strain and some subgroup I-3 strains had the narrowest, growing only at 25° and 30°C. Some spiroplasmas grew well at 41°C, but none grew at 43°C. The ability of spiroplasmas to withstand a wide range of temperatures may reflect the conditions to which they are exposed in nature, including the temperatures of the insect, tick, and/or plant hosts in which they are carried and the plant surfaces from which they may be acquired by arthropods.     

Temperature-dependent physiological and biochemical responses of the marine medaka Oryzias melastigma with consideration of both low and high thermal extremes

This study aimed to investigate temperature effect on physiological and biochemical responses of the marine medaka Oryzias melastigma larvae. The fish were subjected to a stepwise temperature change at a rate of 1 °C/h increasing or decreasing from 25 °C (the control) to six target temperatures (12, 13, 15, 20, 28 and 32 °C) respectively, followed by a 7-day thermal acclimation at each target temperature. The fish were fed ad libitum during the experiment. The results showed that cumulative mortalities were significantly increased at low temperatures (12 and 13 °C) and at the highest temperature (32 °C). For the survivors, their growth profile closely followed the left-skewed ‘thermal performance curve’. Routine oxygen consumption rates of fish larvae were significantly elevated at 32 °C but suppressed at 13 and 15 °C (due to a high mortality, larvae from 12 °C were not examined). Levels of heat shock proteins and activities of malate dehydrogenase and lactate dehydrogenase were also measured in fish larvae exposed at 15, 25 and 32 °C. The activities of both enzymes were significantly increased at both 15 and 32 °C, where the fish larvae probably suffered from thermal discomfort and increased anaerobic components so as to compensate the mismatch of energy demand and supply at these thermal extremes. Coincidently, heat shock proteins were also up-regulated at both 15 and 32 °C, enabling cellular protection. Moreover, the critical thermal maxima and minima of fish larvae increased significantly with increasing acclimation temperature, implying that the fish could develop some degrees of thermal tolerance through temperature acclimation.     

Effect of temperature onin vitro pollen germination in pigeonpea

Pollen of pigeonpea(Cajanus cajan (L.) Millsp.) cultivars H-77-216 and ICPL-151 were cultivatedin vitro at six different temperatures (12, 17, 22, 27, 32, 37 °C). Pollen of cv. H-77-216 started to germinate at 17 °C whereas the pollen of cv. ICPL-151 at 22 °C, the optimal temperatures were 22 and 27 °C, respectively. Pollen germination at different temperatures was found to be positively correlated with the tube length. Per cent pollen bursting increased with rising temperature. The indeterminate cv. H-77-216 showed a wide range of suitable temperatures (17 – 27 °C) for pollen germination while the determinate cv. ICPL-151 had optimum at 27 °C     

FLUCTUATING TEMPERATURE LEADS TO EVOLUTION OF THERMAL GENERALISM AND PREADAPTATION TO NOVEL ENVIRONMENTS

Environmental fluctuations can select for generalism, which is also hypothesized to increase organisms’ ability to invade novel environments. Here, we show that across a range of temperatures, opportunistic bacterial pathogen Serratia marcescens that evolved in fluctuating temperature (daily variation between 24°C and 38°C, mean 31°C) outperforms the strains that evolved in constant temperature (31°C). The growth advantage was also evident in novel environments in the presence of parasitic viruses and predatory protozoans, but less clear in the presence of stressful chemicals. Adaptation to fluctuating temperature also led to reduced virulence in Drosophila melanogaster host, which suggests that generalism can still be costly in terms of reduced fitness in other ecological contexts. While supporting the hypothesis that evolution of generalism is coupled with tolerance to several novel environments, our results also suggest that thermal fluctuations driven by the climate change could affect both species’ invasiveness and virulence.     

THERMAL EVOLUTION OF EGG SIZE IN DROSOPHILA MELANOGASTER

We measured the size of eggs produced by populations of Drosophila melanogaster that had been collected along latitudinal gradients in different continents or that had undergone several years of culture at different temperatures in the laboratory. Australian and South American populations from higher latitudes produced larger eggs when all were compared at a standard temperature. Laboratory populations that had been evolving at 16.5°C produced larger eggs than populations that had evolved at 25°C or 29°C, suggesting that temperature may be an important selective agent in producing the latitudinal clines. Flies from laboratory populations produced larger eggs at an experimental temperature of 16.5°C than at 25°C, and there was no indication of genotype-environment interaction for egg size. Evolution of egg size in response to temperature cannot be accounted for by differences in adult body size between populations. It is not clear which life-history traits are direct targets of thermal selection and which are showing correlated responses, and disentangling these is a task for the future.