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Cited by 827Microbial Metabolic Engineering and BioproductionEvolution and Genetic DynamicsFungal and yeast genetics research

Adaptive laboratory evolution—Principles and applications for biotechnology

Open access
Dragosits, Martin, Mattanovich, Diethard·2013·Microbial Cell Factories
DOI 10.1186/1475-2859-12-64PMC PMC3716822
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Abstract

Adaptive laboratory evolution is a frequent method in biological studies to gain insights into the basic mechanisms of molecular evolution and adaptive changes that accumulate in microbial populations during long term selection under specified growth conditions. Although regularly performed for more than 25 years, the advent of transcript and cheap next-generation sequencing technologies has resulted in many recent studies, which successfully applied this technique in order to engineer microbial cells for biotechnological applications. Adaptive laboratory evolution has some major benefits as compared with classical genetic engineering but also some inherent limitations. However, recent studies show how some of the limitations may be overcome in order to successfully incorporate adaptive laboratory evolution in microbial cell factory design. Over the last two decades important insights into nutrient and stress metabolism of relevant model species were acquired, whereas some other aspects such as niche-specific differences of non-conventional cell factories are not completely understood. Altogether the current status and its future perspectives highlight the importance and potential of adaptive laboratory evolution as approach in biotechnological engineering.

AI snapshot

Written from the paper’s full text.

TL;DR

Adaptive laboratory evolution (ALE) is a long-term selection approach that can improve microbial growth, stress tolerance, and biotechnologically relevant phenotypes, often complementing classical genetic engineering. This review summarizes principles, selection methods, mutation types, trade-offs, and applications across bacteria and yeasts, while noting limitations such as unknown evolved genotypes, trade-offs, and limited data for non-conventional hosts.

Key findings

  • Typical ALE experiments run for about 100–2000 generations over weeks to months; within 100–500 generations, fitness increases of up to 50–100% can be achieved, while an extreme case reached up to 1000% improvement in iron reduction rate for *Geobacter sulfurreducens* after 24 months.
  • Across ALE studies, SNPs accounted for 61% of observed genetic changes, deletions approximately 29%, insertions 7%, and insertion-sequence movements 3%; the Lenski *E. coli* experiment exceeds 50,000 generations.
  • ALE can improve nutrient utilization and stress resistance, but evolved phenotypes often show trade-offs or cross-protection depending on selection conditions; for example, ethanol tolerance in *E. coli* came at the cost of decreased acid resistance, while n-butanol tolerance was weakly compatible with oxidative stress.
  • Large-scale transcriptional rearrangements, gene amplifications, and regulatory mutations are common; epistasis is a major factor, and different evolutionary paths can lead to phenotypic convergence.

Methods

  • This is a review article, not a primary experimental study; it synthesizes principles and applications of ALE for biotechnology.
  • It draws on published ALE experiments with bacteria and yeasts, summarized in Table 1 and Table 2, which list species/strain, selection environment, selection time, and references.
  • The main analysis is a narrative synthesis covering laboratory selection methods, mutation types, nutrient and environmental stress adaptation, evolutionary trade-offs and cross-protection, non-conventional hosts, and future directions.

Limitations

  • The paper states that ALE has inherent limitations: evolved strains have unknown genotypes, risking hitchhiking of unfavorable mutations; selection for one condition often causes trade-offs in others; efficient evolutionary engineering of industrial *S. cerevisiae* may be limited by polyploidy; and computational models cannot yet replace trial-and-error laboratory experiments.
  • The paper states that non-conventional host organisms are understudied, with limited physiological data and niche-specific differences not completely understood.
  • Evident from the text: as a review, it does not perform a systematic search or meta-analysis, and some entries in Tables 1 and 2 report “nd” for selection time, indicating incomplete data in the summarized literature.

Model-written summary. Check it against the paper before relying on it.

Integrity signals

Registry

Crossref · 10.1186/1475-2859-12-64

Cites

189 of 189 references verified0 citation errors · 0 not checkable

Statistics

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Claims the paper makes

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  • Findingcitation

    Based on previous studies, within 100 to 500 generations, corresponding to up to 2 months of selection for a typical E. coli or S. cerevisiae culture, a fitness increase of up to 50-100% can be achieved.

    “Based on previous studies, we can estimate that within 100 to 500 generations (corresponding to up to 2 months of selection for a typical E. coli or S. cerevisiae culture), a fitness increase of up to 50–100% can be achieved.”
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  • Findingown data

    The fitness increase as a function of the total number of generations is not linear.

    “Moreover, it is important to note that the fitness increase as a function of the total number of generations is not linear (Figure 1c).”
    Quote in the paperSubstantial claim
  • Findingnone given

    In recent ALE experiments various types of mutations were detected: single-nucleotide polymorphisms, small scale insertions and deletions, transposable element movements and amplifications as well as deletions of larger genomic regions.

    “In recent ALE experiments various types of mutations were detected: Single-nucleotide polymorphisms (SNPs), small scale insertions and deletions (indels), transposable element (insertion sequence, IS) movements and amplifications as well as deletions of larger genomic regions (Figure 1d).”
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  • Premisenone given

    During direct competition of an ancestral microbial strain and an adapted strain, the increased fitness of the adapted variant will be obvious by its increased frequency in the total population.

    “During direct competition of an ancestral microbial strain and an adapted strain, the increased fitness of the adapted variant will be obvious by its increased frequency in the total population.”
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  • Interpretationreasoning

    It cannot be assumed that a homogenous population is present during any point of a laboratory evolution experiment.

    “Thus, it cannot be assumed that a homogenous population is present during any point of a laboratory evolution experiment.”
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  • May combine two claims. 67

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    Findingown data

    A typical ALE experiment is performed for somewhat between 100 and 2000 generations and usually takes a few weeks up to a few months.

    “In summary Tables 1 and Table 2, it can be seen that a typical ALE experiment is performed for somewhat between 100 and 2000 generations and usually takes a few weeks up to a few months.”
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    Premisecitation

    Biological catalysis offers advantages over chemical synthesis, such as stereo-selective production of chemical compounds.

    “Moreover, biological catalysis offers several advantages over chemical synthesis, such as stereo-selective production of chemical compounds [9].”
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    Systems metabolic engineering has proven very successful for the design and implementation of biotechnological production processes.

    “Synthetic biology is a new emerging field [10,11] and systems metabolic engineering has proven very successful for the design and implementation of biotechnological production processes [12-14].”
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    Premisecitation

    In serial batch cultures for ALE, transfer often occurs before the stationary phase is reached, which is a prerequisite to avoid stationary phase adaptation.

    “As indicated in Figure 1a, in serial batch cultures for ALE, the transfer often occurs before the stationary phase is reached. Clearly, this is a prerequisite to avoid stationary phase adaptation [52].”
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    Premisenone given

    Microbial cells offer important advantages for ALE studies: most have simple nutrient requirements, can be easily cultivated in the laboratory, and generally grow very fast and can be cultivated for several hundred generations within several weeks or months.

    “Microbial cells offer important advantages for ALE studies: (a) most microbial cells have simple nutrient requirements, (b) they can be easily cultivated in the laboratory and (c) microbial cells generally grow very fast and can be cultivated for several hundred generations within several weeks or months”
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    Premisecitation

    Particularly in the last 25 years, there has been an ever increasing number of ALE experiments with Escherichia coli and Saccharomyces cerevisiae being the most prominent organisms under investigation.

    “however, particularly in the last 25 years, there has been an ever increasing number of such experiments with Escherichia coli and Saccharomyces cerevisiae being the most prominent organisms under investigation [26-28].”
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    Premisecitation

    The long-term E. coli adaptation experiment by Lenski's group has exceeded 50000 generations.

    “This single parallel E. coli adaptation experiment is already exceeding 50000 generations [33-35].”
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    Premisecitation

    We cannot clearly predict the scale at which improved phenotypes emerge during laboratory evolution, although recent literature indicates that such predictions are possible in certain growth contexts such as carbon source adaptation.

    “In this context, we cannot clearly predict the scale at which improved phenotypes emerge during laboratory evolution, although recent literature indicates that such predictions are possible in certain growth contexts such as carbon source adaptation [65].”
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    Premisecitation

    Certain environments can lead to fast selection of improved phenotypes, whereas others, including long-term adaptation to environmental pH, represent complex environments where microbial cells show low adaptive potential in terms of absolute fitness gain.

    “Whereas certain environments can lead to fast selection of improved phenotypes, others, including the long-term adaptation to environmental pH [59,63], represent complex environments where microbial cells show low adaptive potential in terms of absolute fitness gain.”
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    Premisecitation

    The fitness increase is usually fast within the first 100 to 500 generations and slows considerably down during the course of ALE.

    “Whereas the fitness increase is usually fast within the first 100 to 500 generations, it slows considerably down during the course of ALE [36].”
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    Premisecitation

    Cells selected under growth limiting conditions can show growth trade-offs in non-limiting conditions and vice versa.

    “In this context, cells selected under growth limiting conditions can show growth trade-offs in non-limiting conditions and vice versa[53].”
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    Premisecitation

    Adaptive laboratory evolution was already performed about a hundred years ago by William Dallinger and during the middle of the last century.

    “Hence, adaptive laboratory evolution was already performed about a hundred years ago by William Dallinger [22] and during the middle of the last century [24,25];”
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    Interpretationreasoning

    The initial choice of conditions, including media composition and limited versus non-limited growth rate, will greatly influence whether the artificially selected microbial host will be suitable for its dedicated application.

    “The initial choice of conditions, including media composition and limited versus non-limited growth rate, will greatly influence whether the artificially selected microbial host will be suitable for its dedicated application.”
    Quote in the paperSubstantial claim

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    Premisecitation

    During adaptive evolution, certain trait values change and are associated with increased Darwinian fitness; an improved phenotype or property is often equal to increased fitness.

    “During adaptive evolution, certain trait values change and are associated with increased (Darwinian) fitness [20]; as such, an improved phenotype or property is often equal to increased fitness.”
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    Premisecitation

    In certain cases ALE can lead to an even higher increase of up to 1000% as shown for the iron reduction rate of Geobacter sulfurreducens, although this increase was only achieved after 24 months of selection.

    “In certain cases ALE can lead to an even higher increase of up to 1000% as recently shown for the iron reduction rate of Geobacter sulfurreducens[123], although this increase was only achieved after 24 months of selection”
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    Premisecitation

    Deletions accounted for approximately 29% of the observed genomic changes, followed by insertions (7%) and IS movements (3%).

    “Deletions accounted for approximately 29% of the observed genomic changes, followed by insertions (7%) and IS movements (3%) [15].”
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    The role of horizontal gene transfer in natural evolution remains a controversial topic.

    “Nevertheless, the role of HGT in natural evolution remains a controversial topic [133].”
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    Premisecitation

    High mutation rates and mutator strains can emerge naturally during laboratory evolution and contribute to the formation of genetic novelty.

    “Interestingly, it was shown that high mutation rates and mutator strains can emerge naturally during laboratory evolution and contribute to the formation of genetic novelty [33,73,139-141].”
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    Premisecitation

    SNPs account for the majority of observed genetic changes, with G:C to T:A mutations being disproportionally frequent.

    “As summarized in a recent review, SNPs (61%, with G:C to T:A mutations being disproportionally frequent) account for the majority of observed genetic changes [15].”
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    Premisecitation

    The toolbox model suggests that prokaryotic metabolic pathways emerge through horizontal gene transfer and are rapidly acquired and lost based on nutrient availability.

    “This so-called toolbox model suggests that prokaryotic metabolic pathways emerge through horizontal gene transfer (HGT) and are rapidly acquired and lost based on nutrient availability [132].”
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    Premisecitation

    Gene duplications are a prerequisite for the development of novel molecular functions.

    “they are a prerequisite for the development of novel molecular functions [129-131].”
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    Premisecitation

    During the first few thousand generations, glucose-evolved E. coli cultures developed larger cell size and a lower final cell density as compared with the ancestor.

    “During the first few thousand generations, glucose-evolved cultures developed larger cell size and a lower final cell density as compared with the ancestor [34].”
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    Premisecitation

    Mutational robustness tends to increase with mutation rate and decline with population size.

    “Furthermore, it was shown that mutational robustness tends to increase with mutation rate and decline with population size [40].”
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    Premisecitation

    In the rpoC study, the biomass yield increased up to 40% and acetate overflow was reduced, whereas the metabolic rate increased and the evolved cells showed a lowered or total loss of motility.

    “In this study, the biomass yield increased up to 40% and acetate overflow was reduced, whereas the metabolic rate increased and the evolved cells showed a lowered or total loss of motility.”
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    Premisecitation

    Microbial metabolic networks often provide appropriate functions at a minimal size, and their complexity seems to be correlated with environmental factors.

    “Microbial metabolic networks often provide appropriate functions at a minimal size [134] and furthermore, their complexity seems to be correlated with environmental factors [135].”
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    Interpretationnone given

    Such mutational adaptations might be counterproductive for some applications of microbial cells in biotechnology.

    “however, such mutational adaptations might be counterproductive for some applications of microbial cells in biotechnology.”
    Quote in the paperNot a substantial claim

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    Premisenone given

    Many ALE studies analyzed the impact and molecular adaptation towards both nutrient and environmental stresses, leading to major findings regarding the emergence of specialists, generalists, the underlying mutations and the applicability of evolved cells for industrial microbiology.

    “Many ALE studies analyzed the impact and molecular adaptation towards both stresses (Table 1 and Table 2), leading to major findings regarding the emergence of specialists, generalists, the underlying mutations and the applicability of evolved cells for industrial microbiology.”
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    Premisecitation

    An experimental E. coli culture could evolve the ability to aerobically utilize citrate, or the ability to use the non-natural carbon source 1,2-propanediol; also, a strain deficient in glutathione biosynthesis was able to develop compensatory mutations to restore this function.

    “For example, an experimental E. coli culture could evolve the ability to aerobically utilize citrate [37], or the ability to use the non-natural carbon source 1,2-propanediol [71]; also, a strain deficient in glutathione biosynthesis was able to develop compensatory mutations to restore this function [147].”
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    Premisecitation

    In a 1997 study, E. coli was evolved in chemostat cultures in order to select for improved growth on glycerol; the authors reported phenotypic changes in terms of colony morphology accompanied with increased growth rate and biomass yield and decreased acetate formation.

    “In a 1997 study, E. coli was evolved in chemostat cultures in order to select for improved growth on glycerol. The authors reported phenotypic changes in terms of colony morphology accompanied with increased growth rate and biomass yield and decreased acetate formation [50].”
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    Premisecitation

    A higher mutation rate can only be beneficial to a certain extent as it also results in a genetic load.

    “It is clear that a higher mutation rate can only be beneficial to a certain extent as it also results in a genetic load [142].”
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    Premisecitation

    In nature as well as in biotechnological processes, microorganisms are challenged with changing environmental conditions, where they experience fluctuations in temperature, pH, oxygenation, atmospheric or hydrostatic pressure but also water and nutrient availability; thus, the selective pressures can be divided in two main categories: nutrient availability and environmental stress.

    “In nature as well as in biotechnological processes, microorganisms are challenged with changing environmental conditions, where they experience fluctuations in temperature, pH, oxygenation, atmospheric or hydrostatic pressure but also water and nutrient availability; thus, the selective pressures can be divided in two main categories: nutrient availability and environmental stress [17].”
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    Premisecitation

    The cumulative number of cell divisions (CCD) was proposed as a more appropriate denominator to evaluate microbial evolution.

    “Only recently, the cumulative number of cell divisions (CCD) was proposed as a more appropriate denominator to evaluate microbial evolution [126].”
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    Premisecitation

    Glucose import, sensing and regulation represent major targets during glucose-limited chemostat growth in S. cerevisiae, and frequent mutations in HXT genes and glucose sensors and regulators such as RGT1 and MIG2 were identified.

    “Glucose import, sensing and regulation represent major targets during glucose-limited chemostat growth, as a further study also identified frequent mutations in HXT genes and glucose sensors and regulators such as RGT1 and MIG2[53].”
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    Premisecitation

    Adaptive changes towards ethanol tolerance in E. coli were linked to the up-regulation of oxyR and nrdR, indicating major changes in the cellular respiratory system and changes in amino acid metabolism.

    “Adaptive changes towards ethanol tolerance were linked to the up-regulation of oxyR and nrdR, indicating major changes in the cellular respiratory system but also changes in amino acid metabolism [85].”
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    Premisecitation

    Transcript analysis in galactose-evolved S. cerevisiae showed up-regulation of trehalose and glycogen metabolism, no mutations in galactose or storage carbohydrate related genes, and mutations in RAS/PKA signaling as a source for increased fitness on galactose medium.

    “Transcript analysis showed up-regulation of trehalose and glycogen metabolism, whereas no mutations were identified in galactose or storage carbohydrate related genes. Interestingly, the authors rather identified mutations in RAS/PKA signaling as a source for increased fitness on galactose medium [106].”
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    Premiseown data

    In xylose-adapted recombinant yeast, no expression changes in HXT genes were identified, although other studies on improved xylose and arabinose utilization indicated that HXT genes may also be involved in increased xylose uptake rates.

    “The authors did not identify expression changes in HXT genes, although other studies on improved xylose and arabinose utilization indicated that HXT genes may also be involved in increased xylose uptake rates [99,104].”
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    Premisecitation

    Growth on glucose was established by genetic and evolutionary engineering in a pyruvate carboxylase-negative S. cerevisiae strain, and a point mutation in the recombinant sfcA gene shifted the co-factor preference from NADH to NADPH.

    “Similarly, Zelle and co-workers established growth on glucose by genetic- and evolutionary engineering in a pyruvate carboxylase-negative S. cerevisiae strain. The authors identified a point mutation in the recombinant sfcA gene, which shifted the co-factor preference from NADH to NADPH [107].”
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    Premisecitation

    Mutations that lead to altered lacZ expression patterns accumulated predominantly in lacI and lacO1 regions of the lac operon, and were responsible for a reduced diauxic lag phase and increased maximum growth rates in lactose growth medium.

    “Mutations that lead to altered expression patterns accumulated predominantly in lacI and lacO1 regions of the lac operon [76]. Altogether, these mutations were responsible for a reduced (diauxic) lag phase and increased maximum growth rates in lactose growth medium.”
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    Premisecitation

    Another study reported the emergence of aneuploidy during glucose-limited growth and amplification of the HXT6 gene with significant divergence between co-evolved S. cerevisiae populations.

    “another study reported the emergence of aneuploidy during glucose-limited growth and amplification of the HXT6 gene with significant divergence between co-evolved populations [94].”
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    Premisecitation

    In evolved ∆pgi E. coli strains, growth rates and glucose uptake rates increased up to 3.6 and up to 2.6 fold as compared with the ancestor, accompanied by different phenotypes regarding acetate and formate production.

    “Growth rates and glucose uptake rates increased up to 3.6 and up to 2.6 fold in evolved ∆pgi strains as compared with the ancestor and were accompanied with different phenotypes regarding acetate and formate production.”
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    Premisecitation

    Early glucose-limited chemostat studies in S. cerevisiae showed that glucose-limited selection led to increased biomass yield and a decreased fermentative capacity.

    “Early studies in glucose-limited chemostats revealed that glucose-limited selection led to increased biomass yield and a decreased fermentative capacity [93,98].”
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    Premisecitation

    In glucose-limited S. cerevisiae, decreased activity levels of enzymes related to glycolysis and correlating changes in expression levels of genes such as ENO1, ENO2, TDH1 and PYK1 and down-regulation of the stress response MSN2/4 were observed.

    “Decreased activity levels of enzymes related to glycolysis but also correlating changes in the expression levels of genes such as ENO1, ENO2, TDH1 and PYK1 and down-regulation of the stress response (MSN2/4) were observed.”
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    Premisecitation

    malT regulatory mutations, although common in aerobic conditions, were absent during oxygen-limited chemostat growth, where mutations in the glucose permease ptsG were common.

    “Strikingly, the authors also noticed that malT regulatory mutations, although common in aerobic conditions, were absent during oxygen-limited chemostat growth, where mutations in the glucose permease ptsG were common [48].”
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    Premisecitation

    The rpoS mutation alone did not positively influence the growth rate in the ancestral ∆pgi E. coli strain; rpoS mutations were epistatically linked to additional mutations.

    “but most importantly the rpoS mutation alone did not positively influence the growth rate in the ancestral ∆pgi strain; rpoS mutations were epistatically linked to additional mutations.”
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    Premisecitation

    In glucose-limited S. cerevisiae, altered cell morphology and higher affinity towards glucose were observed, although no expression change in the major HXT low-affinity glucose transporters was observed.

    “Altered cell morphology was also observed, as well as higher affinity towards glucose, although no expression change in the major HXT low-affinity glucose transporters was observed [98]”
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    Premisecitation

    Mutations in acrAB and marC were found independently in studies of iso- and n-butanol stress in E. coli; further changes were linked to carbon and nitrogen metabolism via gatY and tnaA, enterobactin synthesis, or attenuated rpoS activity via hfq mutations.

    “Mutations in acrAB and marC were found independently in these studies; further changes were linked to carbon and nitrogen metabolism via gatY and tnaA[86], enterobactin synthesis [63] or attenuated rpoS activity via hfq mutations [87].”
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    Premisecitation

    In xylose adaptation studies, increased growth rates and higher ethanol yields were observed in recombinant yeast; trehalose and glycogen metabolic processes were significantly down-regulated, nutrient and stress signaling was affected by down-regulation of YAK1 and CWP1, and major cell wall changes occurred.

    “In xylose adaptation studies, increased growth rates and higher ethanol yields were observed in recombinant yeast. In this case, trehalose and glycogen metabolic processes were significantly down-regulated, nutrient and stress signaling was affected by down-regulation of YAK1 and also accompanied by down-regulation of CWP1 and consequently major cell wall changes [108].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [108] source

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  • May combine two claims.

    Premisecitation

    Evolved ∆pgi E. coli strains had frequent point mutations in rpoS, udhA, pntAB and a prophage deletion; these mutations were not observed in the glucose-adapted wildtype strain, while other regulatory mutations including rpoABC occurred in both backgrounds.

    “The authors found frequent point mutations in rpoS, udhA (soluble transhydrogenase), pntAB and a prophage deletion. They did not observe these mutations in the glucose-adapted wildtype strain; other regulatory mutations (including rpoABC) occurred in both backgrounds”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [70] source

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  • May combine two claims.

    Premisecitation

    Selection for improved galactose utilization could be achieved by serial transfer with a 24% increase of μmax within 62 days, and galactose uptake rates and ethanol production were increased.

    “For other carbon sources, it was shown that selection for improved galactose utilization could be achieved by serial transfer with a 24% increase of μmax within 62 days. Galactose uptake rates and ethanol production were increased.”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [106] source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Experiments on pentose utilization highlighted that increased gene copy numbers of the heterologous genes in recombinant S. cerevisiae can emerge during evolutionary engineering and contribute to increased growth in anaerobic conditions.

    “Furthermore, experiments on pentose utilization highlighted that increased gene copy numbers of the heterologous genes in recombinant S. cerevisiae can emerge during evolutionary engineering and contribute to increased growth in anaerobic conditions [104,109].”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [104] source

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    • cites [109] source

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  • May combine two claims.

    Premisecitation

    In S. cerevisiae, adaptation towards salt tolerance led to increased cell size and increased ploidy; gene expression changes in CTT1 and MSN4 and a high-frequency SNP in the transcriptional regulator MOT2 were observed.

    “In the model yeast S. cerevisiae, adaptation towards salt tolerance led to increased cell size and increased ploidy. Additionally, gene expression changes in CTT1 and MSN4 were observed, as well as a high-frequency SNP in the transcriptional regulator MOT2[116].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [116] source

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  • May combine two claims.

    Premisecitation

    Cross-protection towards lignocellulosic hydrolysates can be evolved in S. cerevisiae; several mutants showed increased fitness in the presence of inhibitor combinations but reduced fitness when exposed to individual hydrolysate components.

    “A recent study showed that cross-protection towards lignocellulosic hydrolysates can be evolved in S. cerevisiae. Interestingly, several mutants showed increased fitness in the presence of inhibitor combinations but reduced fitness when exposed to individual hydrolysate components [112].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [112] source

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  • May combine two claims.

    Premisecitation

    Aerobic chemostat-selected S. cerevisiae cells showed increased fitness in anaerobic glucose-limited chemostats and in acetate-limited chemostats, but decreased fitness in nitrogen-limited batch cultivations.

    “Similarly, aerobic chemostat-selected S. cerevisiae cells also showed increased fitness in anaerobic glucose-limited chemostats and in acetate-limited chemostats, but decreased fitness in nitrogen-limited batch cultivations [53].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [53] source

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  • May combine two claims.

    Interpretationcitation

    Duplications and large scale genomic variations, although they can lead to significant issues such as genomic instability, play a major role in yeast evolution.

    “As such, ALE studies highlight that duplications and large scale genomic variations, although they can lead to significant issues such as genomic instability [152], play a major role in e.g. yeast evolution; this is also indicated by data on wine yeasts, galactose utilization, comparative genomics and drug resistance [115,153-156].”
    Quote in the paperSubstantial claim0 of 6 sources checked
    • cites [152] source

      Not checked in the free run.

    • cites [115] source

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    • cites [153] source

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    • cites [154] source

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    • cites [155] source

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    • cites [156] source

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  • May combine two claims.

    Premisecitation

    A recent study in S. cerevisiae indicated that anticipatory gene regulatory patterns can evolve under cycling salt and oxidative stress conditions on a laboratory time scale within 300 generations, and that cross-stress protection against salt and oxidative stress is strikingly asymmetrical, with oxidative stress protecting against salt stress but not vice versa.

    “In fact, a recent study in S. cerevisiae indicated that anticipatory gene regulatory patterns can evolve under cycling salt and oxidative stress conditions on a laboratory time scale within 300 generations and that cross-stress protection against salt and oxidative stress is strikingly asymmetrical, with oxidative stress protecting against salt stress but not vice versa[120].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [120] source

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  • May combine two claims.

    Interpretationreasoning

    Evolutionary trade-offs and cross-protection against process-relevant parameters are ubiquitous, but the mode of cultivation and the specificities of the selection pressure largely determine to which extent these phenomena arise.

    “Thus, evolutionary trade-offs and cross-protection against process-relevant parameters are ubiquitous, but the mode of cultivation and the specificities of the selection pressure largely determine to which extent these phenomena arise.”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Increased copper tolerance in S. cerevisiae was reported to be mediated by a genomic amplification of CUP1, decreased basal levels of the copper transporters CTR2 and CCC2, and decreased activity of antioxidant enzymes.

    “Increased copper tolerance was reported to be mediated by a genomic amplification of CUP1, decreased basal levels of the copper transporters CTR2 and CCC2 and decreased activity of antioxidant enzymes [117,128].”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [117] source

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    • cites [128] source

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  • May combine two claims.

    Interpretationnone given

    Epistatic interactions can be hardly inferred from metabolic modeling studies, although significant epistatic connections for strain engineering may be easily identified in ALE studies.

    “Importantly, epistatic interactions can be hardly inferred from metabolic modeling studies although significant epistatic connections for strain engineering may be easily identified in ALE studies.”
    Quote in the paperNot a substantial claim

  • May combine two claims.

    Premisecitation

    Increased ethanol tolerance in S. cerevisiae was mediated by mutations in the translational regulator SSD1 and UTH1, a protein of unknown function, and indicated that cell wall stability is a major factor in ethanol tolerance of S. cerevisiae.

    “In another study, increased ethanol tolerance was mediated by mutations in the translational regulator SSD1 and UTH1, a protein of unknown function, and indicated that cell wall stability is a major factor in ethanol tolerance of S. cerevisiae[119].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [119] source

      Not checked in the free run.

  • May combine two claims.

    Interpretationreasoning

    Long-term adaptation leads to both large-scale transcriptional de-regulation and optimization of terminal nodes (fine-tuning of the expression levels of individual genes).

    “From these data it can be concluded that: (a) long-term adaptation leads to both large-scale transcriptional de-regulation and optimization of terminal nodes (fine-tuning of the expression levels of individual genes)”
    Quote in the paperSubstantial claim

  • May combine two claims.

    Premisecitation

    Glycerol-limited growth led to stable mutants in a single E. coli population; one mutant showed increased growth in glycerol-sufficient batch cultures and a second mutant did not; the mutant with increased batch growth also showed increased fitness under general nutrient limitation, heat and osmotic stress.

    “Glycerol-limited growth led to stable mutants in a single E. coli population, where one mutant showed increased growth in glycerol-sufficient batch cultures and a second mutant did not. Furthermore, the mutant that showed increased growth in batch culture also showed increased fitness under general nutrient limitation, heat and osmotic stress [50].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [50] source

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  • May combine two claims.

    Premisecitation

    Multi-stress resistant S. cerevisiae populations can be evolved; batch selection for freeze-thaw tolerance also selected for increased thermal, oxidative and ethanol stress resistance.

    “Another interesting study analyzed to which extent multi-stress resistant S. cerevisiae populations can be evolved. The authors tested various chemostat and batch stress regimes and found that batch selection for freeze-thaw tolerance also selected for increased thermal, oxidative and ethanol stress resistance [113].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [113] source

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  • May combine two claims.

    Premisecitation

    Yeast cells that were evolved for efficient galactose utilization subsequently showed trade-offs when grown on glucose as carbon source; this effect was due to antagonistic pleiotropy based on mutations of the regulatory protein RAS2.

    “Yeast cells that were evolved for efficient galactose utilization [106] subsequently showed trade-offs when grown on glucose as carbon source. This effect was due to antagonistic pleiotropy that was based on mutations of the regulatory protein RAS2[162].”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [106] source

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    • cites [162] source

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  • May combine two claims.

    Premisecitation

    Core sub-networks and pathways tend to be well conserved, but the up- and downstream connections to other cellular networks vary significantly among different species and lead to species-specific differences of stress resistance in adverse environments.

    “Whereas core sub-networks and pathways tend to be well conserved, the up- and downstream connections to other cellular networks vary significantly among different species and lead to species-specific differences of stress resistance in adverse environments [169,170]”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [169]

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    • cites [170]

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  • May combine two claims.

    Premisecitation

    In S. cerevisiae, cobalt resistance also improved tolerance for other metal ions, and pulsed cobalt stress exposure during laboratory evolution resulted in evolutionary cross-protection to thermal and oxidative stress.

    “Examples for adaptive cross-protection are also available for S. cerevisiae where cobalt resistance also improved tolerance for other metal ions and pulsed cobalt stress exposure during laboratory evolution even resulted in evolutionary cross-protection to thermal and oxidative stress [114].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [114] source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    n-butanol tolerance in E. coli is weakly compatible with oxidative stress and leads to trade-offs in hexane and chloramphenicol resistance.

    “and n-butanol tolerance is weakly compatible with oxidative stress [63] and leads to trade-offs in hexane and chloramphenicol resistance [86].”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [63] source

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    • cites [86] source

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  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    For eukaryotic host organisms such as yeasts, alternating asexual and sexual propagation during selection can be applied in order to decouple beneficial and unsound mutations and thus reduce potential deficits of laboratory evolution experiments.

    “Moreover, for eukaryotic host organisms such as yeasts, alternating asexual and sexual propagation during selection can be applied in order to decouple beneficial and unsound mutations and thus reduce potential deficits of laboratory evolution experiments [118].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [118] source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S2 (checkable). May combine two claims.

    Interpretationreasoning

    Computational approaches provide important benefits towards rational engineering of microbial cell factories but cannot replace or prevent 'trial and error' laboratory experiments for the foreseeable future.

    “In this context, computational approaches provide important benefits towards rational engineering of microbial cell factories but as it can be foreseen right now, they cannot replace or prevent ‘trial and error’ laboratory experiments for the foreseeable future.”
    Quote in the paperNot a substantial claim
    • Rests on an assumption: Enormous computational power is necessary to simulate living cells, even on a non-evolutionary scale.
    • Rests on an assumption: Even for well-studied organisms such as E. coli and S. cerevisiae, approximately 30% of genes have unknown function.
    • Rests on an assumption: The complex interactions of living organisms with their environment are not well understood.
    • Rests on an assumption: Our understanding of the effect of many environmental stressors on cells, their precise action on certain cellular components, the mode of stress sensing, cellular signal transduction and cross-talk between signal transduction pathways is still limited.

  • Probably not a substantial claim: may not pass S3 (load-bearing). May combine two claims.

    Premisecitation

    Synthetic biology is highly structured and biological networks are split into smaller functional modules, leading to complex optimization needs.

    “In contrast to classical approaches it is highly structured and biological networks are split into smaller functional modules [11,183], leading to complex optimization needs.”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [11] source

      Not checked in the free run.

    • cites [183]

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Full-scale models of microorganisms are available and microbial growth can be simulated.

    “Full-scale models of microorganisms are available [185,186] and we can simulate microbial growth.”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [185]

      Not checked in the free run.

    • cites [186]

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  • May combine two claims.

    Premisecitation

    A recent study with S. cerevisiae and Saccharomyces paradoxus showed that the evolutionary rescue frequency was positively correlated with stress concentration during 100 generations of evolution in S. cerevisiae but negatively correlated in S. paradoxus.

    “Towards this end, a recent study with S. cerevisiae and Saccharomyces paradoxus showed that the evolutionary rescue (ER) frequency was positively correlated with stress concentration during 100 generations of evolution in S. cerevisiae but negatively correlated in S. paradoxus[44].”
    Quote in the paperSubstantial claim0 of 1 sources checked
    • cites [44] source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Other studies used ethylmethanesulfonate (EMS) treated Bacillus boroniphilus to select for increased boron resistance or EMS-treated S. cerevisiae in order to achieve increased nutrient utilization and stress resistance.

    “Other studies used ethylmethanesulfonate (EMS) treated Bacillus boroniphilus to select for increased boron resistance [88] or EMS-treated S. cerevisiae in order to achieve increased nutrient utilization and stress resistance [95,113,114].”
    Quote in the paperNot a substantial claim0 of 4 sources checked
    • cites [88] source

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    • cites [95] source

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    • cites [113] source

      Not checked in the free run.

    • cites [114] source

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  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    In silico approaches are widely applied in order to address evolutionary questions.

    “Similarly, in silico approaches are widely applied in order to address evolutionary questions [40,172].”
    Quote in the paperNot a substantial claim0 of 2 sources checked
    • cites [40] source

      Not checked in the free run.

    • cites [172]

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  • May combine two claims.

    Premisecitation

    Microbial gene regulatory networks are generally very robust towards perturbations and large-scale re-wiring results in a significant number of viable clones with increased evolvability as compared with the wildtype.

    “Microbial gene regulatory networks are generally very robust towards perturbations and large-scale re-wiring results in a significant number of viable clones with increased evolvability as compared with the wildtype [181].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [181]

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Computational models are well-suited to study evolutionary change on the population level, whereas in silico evolution of artificial cells that include simulated regulatory networks is currently very limited.

    “Computational models are well-suited to study evolutionary change on the population level, whereas in silico evolution of artificial cells that include simulated regulatory networks are currently very limited [21].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [21] source

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Certain functions such as overflow metabolism in S. cerevisiae may have evolved in order to provide an evolutionary advantage in specific environments and to protect carbon resources from other competing species.

    “Certain functions such as overflow metabolism in S. cerevisiae may have evolved in order to provide an evolutionary advantage in specific environments and to protect carbon resources from other competing species [175].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [175]

      Not checked in the free run.

  • May combine two claims.

    Premisecitation

    Laboratory selection can lead to large-scale rewiring of gene regulatory networks, but an approach called gTME, global transcription machinery engineering, can also yield similar results.

    “For example, laboratory selection can lead to large-scale rewiring of gene regulatory networks, but an approach, that the authors called gTME, global transcription machinery engineering, can also yield similar results [2].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [2] source

      Not checked in the free run.

  • Probably not a substantial claim: may not pass S3 (load-bearing).

    Premisecitation

    Co-cultivation of multiple organisms can be exploited in order to increase antibiotic production, such as in the case of co-cultivation of Streptomyces clavuligerus and Staphylococcus aureus.

    “Besides insights into co-evolutionary behavior, this approach can be exploited in order to increase antibiotic production such as in the case of co-cultivation of Streptomyces clavuligerus and Staphylococcus aureus[55].”
    Quote in the paperNot a substantial claim0 of 1 sources checked
    • cites [55] source

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78 of 78

  • The paper relies on this without support.

    Mutations are the basis underlying genetic change and the selection of improved phenotypes in nature as well as in the laboratory.

    “Mutations are the basis underlying genetic change and the selection of improved phenotypes in nature as well as in the laboratory.”

  • The paper relies on this without support.

    Microbial cells evolved mechanisms to successfully cope with stressful conditions in order to maintain cellular homeostasis.

    “Microbial cells evolved mechanisms to successfully cope with such stressful conditions in order to maintain cellular homeostasis.”

  • The paper relies on this without support.

    Efficiency of nutrient source utilization is an important aspect of microbial growth and for industrial purposes.

    “Efficiency of nutrient source utilization is an important aspect of microbial growth and for industrial purposes it may be governed by factors such as substrate cost or increased bioconversion rates.”

  • The paper relies on this without support.

    In industrial processes microbial cells may be challenged with non-natural stresses that may result in poor and non-competitive process performance.

    “In contrast to natural environments, in industrial processes microbial cells may be challenged with non-natural stresses that may result in poor and non-competitive process performance.”

  • The paper relies on this without support.

    The total number of generations is commonly used to estimate the emergence of adaptive mutations.

    “the total number of generations is commonly used to estimate the emergence of adaptive mutations.”

  • The paper relies on this without support.

    The evolution of novel functions and pathways can be hardly observed in a laboratory evolution experiment, even with microbial cells and their fast growth rates.

    “The evolution of novel functions and pathways can be hardly observed in a laboratory evolution experiment, even with microbial cells and their fast growth rates.”

  • The paper relies on this without support.

    rpoS plays a crucial role towards survival in high osmotic conditions.

    “Furthermore, it is known that rpoS plays a crucial role towards survival in high osmotic conditions.”

  • The paper relies on this without support.

    Mesophilic organisms such as E. coli and S. cerevisiae have inherent properties that limit their use in certain applications, e.g. high temperature processes.

    “Mesophilic organisms such as E. coli and S. cerevisiae have inherent properties that limit their use in certain applications, e.g. high temperature processes.”

  • The paper relies on this without support.

    Limited data on the physiology of non-conventional organisms still represent a significant obstacle towards rational host cell engineering.

    “Currently, limited data on the physiology of such organisms still represent a significant obstacle towards rational host cell engineering.”

  • The paper relies on this without support.

    Enormous computational power is necessary to simulate living cells, even on a non-evolutionary scale.

    “Enormous computational power is necessary to simulate living cells, even on a non-evolutionary scale.”

  • The paper relies on this without support.

    Even for well-studied organisms such as E. coli and S. cerevisiae, approximately 30% of genes have unknown function.

    “Additionally, even for well-studied organisms such as E. coli and S. cerevisiae, there is still a significant number (approximately 30%) of genes with unknown function.”

  • The paper relies on this without support.

    The complex interactions of living organisms with their environment are not well understood.

    “Furthermore, the complex interactions of living organisms with their environment are not well understood.”

  • The paper relies on this without support.

    Our understanding of the effect of many environmental stressors on cells, their precise action on certain cellular components, the mode of stress sensing, cellular signal transduction and cross-talk between signal transduction pathways is still limited.

    “Specifically, our understanding of the effect of many environmental stressors on cells, their precise action on certain cellular components, the mode of stress sensing, cellular signal transduction and cross-talk between signal transduction pathways is still limited.”

  • The authors state this as an assumption.

    Evolution is constrained to work on available blueprints.

    “As François Jacob pointed out: “Evolution is a tinkerer”, and as such evolution is constrained to work on available blueprints.”

  • The paper relies on this without support.

    Genetic engineers have the possibility to rapidly construct novel circuits.

    “However, genetic engineers have the possibility to rapidly construct novel circuits.”
15 of 15

Claims other papers make about this one

Each is checked against this paper’s own text: does it say what the citing paper says it does?

  • Adaptive evolutionary engineering based on metabolic engineering can allow microorganisms to quickly obtain excellent phenotypes, but there are problems such as unclear gene targets and negative mutation interference.

    “Adaptive evolutionary engineering based on metabolic engineering can allow microorganisms to quickly obtain excellent phenotypes, but there are problems such as unclear gene targets and negative mutation interference [90,91].”

    From Recent Developments in Lignocellulosic Biofuels, a Renewable Source of Bioenergy (2022) · cites this paper as [90]

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Authors
Dragosits, M., & Mattanovich, D.
Affiliations
BOKU University (AT)
Corresponding author
Dragosits, Martin
ORCID
Venue
Microbial Cell Factories
Publisher
Springer Science and Business Media LLC
Published
2013
Volume
12
Issue
1
Pages
64
ISSN
1475-2859 (ISSN-L)
DOI
10.1186/1475-2859-12-64
PMC
PMC3716822
OpenAlex
W2115941371
Semantic Scholar
09e8fffffe29d6b53bee072e4f632e32239a43f8
Cited by
827
References
188
Citations by year
2026: 80 · 2025: 85 · 2024: 99 · 2023: 69 · 2022: 63 · 2021: 80 · 2020: 84 · 2019: 67 · 2018: 66 · 2017: 52 · 2016: 36 · 2015: 23 · 2014: 21 · 2013: 2
FWCI
2.57
Type
journal article
Language
en
Open access
Licence
cc-by
Locations
  • Microbial Cell Factories (publishedVersion, cc-by)
  • Europe PMC (PubMed Central) (publishedVersion, cc-by)
  • PubMed Central (publishedVersion, cc-by)
  • PubMed (publishedVersion)
  • BOK:ePub (Universitätsbibliothek der Universität f Bodenkultur Wien) (submittedVersion)
Topics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Microbial Metabolic Engineering and Bioproduction
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Genetics › Evolution and Genetic Dynamics
  • Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Fungal and yeast genetics research
Keywords
adaptive laboratory evolution, microbial engineering, molecular evolution, nutrient metabolism, stress metabolism, comparative genomics, transcriptomics, next-generation sequencing
MeSH terms
Biotechnology, Evolution, Molecular, Gene Regulatory Networks

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