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The Caenorhabditis elegans Lifespan Machine

Nature Methods volume 10, pages 665–670 (2013)Cite this article

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Abstract

The measurement of lifespan pervades aging research. Because lifespan results from complex interactions between genetic, environmental and stochastic factors, it varies widely even among isogenic individuals. The actions of molecular mechanisms on lifespan are therefore visible only through their statistical effects on populations. Indeed, survival assays in Caenorhabditis elegans have provided critical insights into evolutionarily conserved determinants of aging. To enable the rapid acquisition of survival curves at an arbitrary statistical resolution, we developed a scalable imaging and analysis platform to observe nematodes over multiple weeks across square meters of agar surface at 8-μm resolution. The automated method generates a permanent visual record of individual deaths from which survival curves are constructed and validated, producing data consistent with results from the manual method of survival curve acquisition for several mutants in both standard and stressful environments. Our approach permits rapid, detailed reverse-genetic and chemical screens for effects on survival and enables quantitative investigations into the statistical structure of aging.

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Figure 1: Lifespan Machine workflow.
Figure 2: Automated wild-type survival data.
Figure 3: Automated mutant survival data.
Figure 4: Automated stress-resistance assays.

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Acknowledgements

We thank J. Alcedo (Wayne State University) for providing the hsf-1 and glp-1 mutant strains, X. Manière (Université Paris Descartes) for providing the NEC937 strain, B. Ward and D. Marks for critical reading of our manuscript and C. Romero, D. Marks and the members of the Fontana lab for helpful discussions and encouragement throughout this project. We thank T. Kolokotrones, E. Smith and L.J. Wei for discussions and statistical advice and M. Miranda, our departmental IT specialist, for patiently meeting our needs for data storage. Some nematode strains used in this work were provided by the Caenorhabditis Genetics Center, which is funded by the National Center for Research Resources. This work was funded by the US National Institutes of Health through grants R03 AG032481, R03 AG032481-S1 and R01 AG034994.

Author information

Author notes

  1. Javier Apfeld and Walter Fontana: These authors contributed equally to this work.

Authors and Affiliations

  1. Department of Systems Biology, Harvard Medical School, Boston, Massachusetts, USA

    Nicholas Stroustrup, Bryne E Ulmschneider, Zachary M Nash, Isaac F López-Moyado, Javier Apfeld & Walter Fontana

Authors
  1. Nicholas Stroustrup
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  2. Bryne E Ulmschneider
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  3. Zachary M Nash
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Contributions

N.S. designed and implemented hardware and software. N.S. and B.E.U. constructed and calibrated equipment. N.S. and J.A. conceived and designed experiments. N.S., B.E.U., J.A., Z.M.N. and I.F.L.-M. performed experiments. N.S. designed analytic tools. N.S., J.A. and W.F. provided guidance, analyzed data, interpreted results and wrote the manuscript. J.A and W.F. are co-last authors.

Corresponding authors

Correspondence to Nicholas Stroustrup, Javier Apfeld or Walter Fontana.

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Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Notes 1–16 (PDF 6347 kb)

Supplementary Data

Directory containing all lifespan data referenced in the manuscript (ZIP 277 kb)

Supplementary Table 1

Population sizes and statistical survival data (XLS 82 kb)

Raw time lapse of a single plate with wild-type animals

A time-lapse video covering 3 weeks of automatic image captures by the Lifespan Machine of a single plate with wild-type animals. (MOV 1293 kb)

Annotated time lapse of a single plate with wild-type animals

A time-lapse video covering 3 weeks of automatic image captures by the Lifespan Machine of a single plate with wild-type animals (Supplementary Video 1), overlaid with metadata from image analysis. Animals are colored according to their movement class. Animals that manifest locomotion are colored purple. Stationary animals that manifest posture changes are colored yellow. Completely motionless (dead) animals are colored red. Blue objects have been excluded as nonworm objects during the validation step. The survival curve of the plate population is shown in yellow, and the aggregated survival curve for the population across all plates on the entire scanner is shown in red. (MOV 1377 kb)

Raw time lapse of a single plate with age-1(hx546) mutants

A time-lapse video covering 3 weeks of automatic image captures by the Lifespan Machine of a single plate with age-1(hx546) mutants. (MOV 1812 kb)

Annotated time lapse of a single plate with age-1(hx546) mutants

A time-lapse video covering 3 weeks of automatic image captures by the Lifespan Machine of a single plate with age-1(hx546) mutants, overlaid with metadata from image analysis. The coloring is as described in Supplementary Video 2. (MOV 2346 kb)

Annotated time lapse of a single plate with unc-64(e246) mutants

A time-lapse video covering 3 weeks of automatic image captures by the Lifespan Machine of a single plate with unc-64(e246) mutants, overlaid with metadata from image analysis. The coloring is as described in Supplementary Video 2. (MOV 935 kb)

Worm Browser demo

00:03 The Worm Browser is a desktop utility that allows users to interact with Lifespan Machine data. 00:08 Data from different experiments can be easily accessed. 00:21 Worm deaths can be inspected in a variety of ways. For example, all animals observed in an experiment can be sorted by their death time, or a single plate can be observed in isolation. Individuals can be viewed at their time of death or at the time of death of the last animal to die in the experiment. 00:26 Objects that have been incorrectly classified as worms can be flagged for exclusion. 00:33 The death transition of a worm can be inspected by clicking through a time-lapse video of late-life movements. The first (upper) colored bar underneath the image of the worm represents the time during which the animal is detected as having ceased locomotion. The switch from yellow to red marks the worm's transition from changing posture to being completely motionless and is taken as the time of death. 00:33 The second (lower) colored bar is used for human annotation of the animal's death time. These data are stored and later used to generate various diagnostics. 01:10 In a number of cases, several worms may die closely juxtaposed. These cases can be annotated and taken into account when the survival curves are assembled. 01:27 The Worm Brower generates a variety of statistical outputs, including survival curves, quantification of worm movement and comparisons between automated and manual annotations. (AVI 15642 kb)

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Stroustrup, N., Ulmschneider, B., Nash, Z. et al. The Caenorhabditis elegans Lifespan Machine. Nat Methods 10, 665–670 (2013). https://doi.org/10.1038/nmeth.2475

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