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Application of a mathematical model to describe the effects of chlorpyrifos on Caenorhabditis elegans development.

Boyd WA, Smith MV, Kissling GE, Rice JR, Snyder DW, Portier CJ, Freedman JH - PLoS ONE (2009)

Bottom Line: Concentration response curves with respect to several model-estimated quantities (numbers of measured nematodes, mean log(TOF) and log(EXT), growth rates, and time to reach change points) showed a significant decrease in C. elegans growth with increasing chlorpyrifos concentration.Statistical tests confirmed a significant concentration effect on several model endpoints.The most noticeable effect on growth occurred during early larval stages: L2 and L3.

View Article: PubMed Central - PubMed

Affiliation: Biomoleclular Screening Branch, National Toxicology Program, Research Triangle Park, North Carolina, USA.

ABSTRACT

Background: The nematode Caenorhabditis elegans is being assessed as an alternative model organism as part of an interagency effort to develop better means to test potentially toxic substances. As part of this effort, assays that use the COPAS Biosort flow sorting technology to record optical measurements (time of flight (TOF) and extinction (EXT)) of individual nematodes under various chemical exposure conditions are being developed. A mathematical model has been created that uses Biosort data to quantitatively and qualitatively describe C. elegans growth, and link changes in growth rates to biological events. Chlorpyrifos, an organophosphate pesticide known to cause developmental delays and malformations in mammals, was used as a model toxicant to test the applicability of the growth model for in vivo toxicological testing.

Methodology/principal findings: L1 larval nematodes were exposed to a range of sub-lethal chlorpyrifos concentrations (0-75 microM) and measured every 12 h. In the absence of toxicant, C. elegans matured from L1s to gravid adults by 60 h. A mathematical model was used to estimate nematode size distributions at various times. Mathematical modeling of the distributions allowed the number of measured nematodes and log(EXT) and log(TOF) growth rates to be estimated. The model revealed three distinct growth phases. The points at which estimated growth rates changed (change points) were constant across the ten chlorpyrifos concentrations. Concentration response curves with respect to several model-estimated quantities (numbers of measured nematodes, mean log(TOF) and log(EXT), growth rates, and time to reach change points) showed a significant decrease in C. elegans growth with increasing chlorpyrifos concentration.

Conclusions: Effects of chlorpyrifos on C. elegans growth and development were mathematically modeled. Statistical tests confirmed a significant concentration effect on several model endpoints. This confirmed that chlorpyrifos affects C. elegans development in a concentration dependent manner. The most noticeable effect on growth occurred during early larval stages: L2 and L3. This study supports the utility of the C. elegans growth assay and mathematical modeling in determining the effects of potentially toxic substances in an alternative model organism using high-throughput technologies.

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C. elegans growth from L1 to adult.Nematode frequency distributions of log(EXT) values for control nematode cohorts sampled at loading 0 h (brown) and following 12 (green), 24 (light blue), 36 (purple), 48 (black), 60 (dark blue), and 72 (red) h incubations. Large modes to the left of the loaded nematodes (log(EXT) ≈2–3) indicate the second generation of embryos corresponding to 60 and 72 h cohorts. Vertical lines at log(EXT)  = 3.83 and 6.04 divide the growth response into three sections: initial growth from starved L1s, larval growth from L2 to L4, and adult growth, respectively.
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pone-0007024-g001: C. elegans growth from L1 to adult.Nematode frequency distributions of log(EXT) values for control nematode cohorts sampled at loading 0 h (brown) and following 12 (green), 24 (light blue), 36 (purple), 48 (black), 60 (dark blue), and 72 (red) h incubations. Large modes to the left of the loaded nematodes (log(EXT) ≈2–3) indicate the second generation of embryos corresponding to 60 and 72 h cohorts. Vertical lines at log(EXT)  = 3.83 and 6.04 divide the growth response into three sections: initial growth from starved L1s, larval growth from L2 to L4, and adult growth, respectively.

Mentions: Distributions of log(EXT) at each of the six time points for the control cohorts are presented in Figure 1. These distributions shifted to the right with increasing incubation time; L1s loaded at t = 0 h had a peak frequency at log(EXT) value near 3.1, while nematodes observed following 60 h and 72 h incubations had peak frequency at log(EXT) values near 6.3. Optical density, as measured by EXT, tends to increase as nematodes mature [10]. Microscopic observations showed that L1 nematodes matured to L2s by 12 h, L3s by 24 h, L4s by 48 h, and adults by 60 h. After 60 h, adults showed a small increase in optical density but mainly produced offspring. The large numbers of measurements with log(EXT) values between 2 to 3.4 observed at 60 h and 72 h included unhatched embryos laid by the adults and the next generation of L1 larvae.


Application of a mathematical model to describe the effects of chlorpyrifos on Caenorhabditis elegans development.

Boyd WA, Smith MV, Kissling GE, Rice JR, Snyder DW, Portier CJ, Freedman JH - PLoS ONE (2009)

C. elegans growth from L1 to adult.Nematode frequency distributions of log(EXT) values for control nematode cohorts sampled at loading 0 h (brown) and following 12 (green), 24 (light blue), 36 (purple), 48 (black), 60 (dark blue), and 72 (red) h incubations. Large modes to the left of the loaded nematodes (log(EXT) ≈2–3) indicate the second generation of embryos corresponding to 60 and 72 h cohorts. Vertical lines at log(EXT)  = 3.83 and 6.04 divide the growth response into three sections: initial growth from starved L1s, larval growth from L2 to L4, and adult growth, respectively.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2737145&req=5

pone-0007024-g001: C. elegans growth from L1 to adult.Nematode frequency distributions of log(EXT) values for control nematode cohorts sampled at loading 0 h (brown) and following 12 (green), 24 (light blue), 36 (purple), 48 (black), 60 (dark blue), and 72 (red) h incubations. Large modes to the left of the loaded nematodes (log(EXT) ≈2–3) indicate the second generation of embryos corresponding to 60 and 72 h cohorts. Vertical lines at log(EXT)  = 3.83 and 6.04 divide the growth response into three sections: initial growth from starved L1s, larval growth from L2 to L4, and adult growth, respectively.
Mentions: Distributions of log(EXT) at each of the six time points for the control cohorts are presented in Figure 1. These distributions shifted to the right with increasing incubation time; L1s loaded at t = 0 h had a peak frequency at log(EXT) value near 3.1, while nematodes observed following 60 h and 72 h incubations had peak frequency at log(EXT) values near 6.3. Optical density, as measured by EXT, tends to increase as nematodes mature [10]. Microscopic observations showed that L1 nematodes matured to L2s by 12 h, L3s by 24 h, L4s by 48 h, and adults by 60 h. After 60 h, adults showed a small increase in optical density but mainly produced offspring. The large numbers of measurements with log(EXT) values between 2 to 3.4 observed at 60 h and 72 h included unhatched embryos laid by the adults and the next generation of L1 larvae.

Bottom Line: Concentration response curves with respect to several model-estimated quantities (numbers of measured nematodes, mean log(TOF) and log(EXT), growth rates, and time to reach change points) showed a significant decrease in C. elegans growth with increasing chlorpyrifos concentration.Statistical tests confirmed a significant concentration effect on several model endpoints.The most noticeable effect on growth occurred during early larval stages: L2 and L3.

View Article: PubMed Central - PubMed

Affiliation: Biomoleclular Screening Branch, National Toxicology Program, Research Triangle Park, North Carolina, USA.

ABSTRACT

Background: The nematode Caenorhabditis elegans is being assessed as an alternative model organism as part of an interagency effort to develop better means to test potentially toxic substances. As part of this effort, assays that use the COPAS Biosort flow sorting technology to record optical measurements (time of flight (TOF) and extinction (EXT)) of individual nematodes under various chemical exposure conditions are being developed. A mathematical model has been created that uses Biosort data to quantitatively and qualitatively describe C. elegans growth, and link changes in growth rates to biological events. Chlorpyrifos, an organophosphate pesticide known to cause developmental delays and malformations in mammals, was used as a model toxicant to test the applicability of the growth model for in vivo toxicological testing.

Methodology/principal findings: L1 larval nematodes were exposed to a range of sub-lethal chlorpyrifos concentrations (0-75 microM) and measured every 12 h. In the absence of toxicant, C. elegans matured from L1s to gravid adults by 60 h. A mathematical model was used to estimate nematode size distributions at various times. Mathematical modeling of the distributions allowed the number of measured nematodes and log(EXT) and log(TOF) growth rates to be estimated. The model revealed three distinct growth phases. The points at which estimated growth rates changed (change points) were constant across the ten chlorpyrifos concentrations. Concentration response curves with respect to several model-estimated quantities (numbers of measured nematodes, mean log(TOF) and log(EXT), growth rates, and time to reach change points) showed a significant decrease in C. elegans growth with increasing chlorpyrifos concentration.

Conclusions: Effects of chlorpyrifos on C. elegans growth and development were mathematically modeled. Statistical tests confirmed a significant concentration effect on several model endpoints. This confirmed that chlorpyrifos affects C. elegans development in a concentration dependent manner. The most noticeable effect on growth occurred during early larval stages: L2 and L3. This study supports the utility of the C. elegans growth assay and mathematical modeling in determining the effects of potentially toxic substances in an alternative model organism using high-throughput technologies.

Show MeSH
Related in: MedlinePlus