Neuroimaging results altered by varying analysis pipelines

Seventy laboratories that analysed the same neuroimaging data each produced different results. This finding highlights the potential consequences of a lack of standardized pipelines for processing complex data.
Martin Lindquist is in the Department of Biostatistics, Johns Hopkins Bloomberg School of Public Health, Baltimore, Maryland 21205, USA.

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For most types of big data, from genome sequences to medical images, there is no single ‘best’ way to process the data. This issue is exemplified by the substantial differences in how individual laboratories preprocess and analyse data from functional magnetic resonance imaging (fMRI) experiments, which generate information about brain activity. Indeed, a survey of fMRI studies found that nearly every study used a different analysis pipeline1. Writing in Nature, Botvinik-Nezer et al.2 provide further evidence of this variability, highlighting how analytical choices made by individual researchers can greatly influence the findings gleaned from an fMRI data set. The work is bound to spark lively discussion.

Functional MRI experiments produce a series of images of the brain at work. These images go through several stages of processing and analysis to determine which brain regions show significant activity. However, the choice of pipeline can alter the outcome of an fMRI study. In 2012, for instance, 6,912 unique processing and analysis pipelines were applied to the same fMRI data set3. Many of the pipelines indicated that the same general brain regions were active, but the locations at which neural activity was deemed to be highest varied widely depending on the pipeline used.

Botvinik-Nezer et al. explored this phenomenon further. The authors gave 70 independent research teams the same fMRI data set, generated from 108 people performing decision-related tasks. The teams were asked to use the data set to test nine hypotheses, each of which posited that activity in a specific brain region related to a particular feature of the tasks. This allowed Botvinik-Nezer et al. to evaluate the impact of analytical flexibility on fMRI results ‘in the wild’ (rather than performing the analyses themselves, as was done in the 2012 study). Notably, no two teams chose identical workflows to analyse the data, resulting in substantial variation in the results (Fig. 1).

Figure 1 | Implications of choosing a neuroimaging pipeline. Botvinik-Nezer et al.2 report that researchers process neuroimaging data using a wide variety of pipelines, which can produce varying results. In this simplified example, the pipeline has three steps: spatial smoothing of the images to reduce noise, which in this example is done to three different degrees; statistical modelling, which in this example can be performed in one of two ways; and ‘thresholding’ of statistical tests associated with these models to determine the level at which neuronal activity in each brain region is deemed to be significant, which in this example is set to two different values. Making different choices for each step leads to a different end point — the red dots represent how activation moves throughout the brain depending on which pipeline is used.

It is standard in neuroimaging to test types of hypothesis such as the nine put forward by Botvinik-Nezer et al. by generating a statistical map. The map comprises a compendium of statistical tests performed on different parts (voxels) of the image. The results of these tests are subjected to a process called thresholding to set the level, for each voxel, at which activity picked up by the experiment is deemed to indicate real neuronal activity rather than noise. The authors found considerable differences between each research team’s results even when the underlying statistical maps they had used were highly correlated.

The strongest factor in explaining the differences between each team’s results was the spatial smoothness of the data being analysed. Spatial smoothing is a preprocessing step in which the activity of each voxel is averaged with that of its neighbours — a process designed to reduce noise. Higher estimated smoothness was associated with a greater likelihood of reaching a significant outcome for each hypothesis.

The fact that each team’s results were so pipeline-dependent is highly problematic, particularly because the exact configuration of analytical pipelines is often poorly described in research articles. Moreover, sensitivity analyses — which assess how different pipeline choices might affect an experiment’s outcome — are rarely performed in neuroimaging. However, Botvinik-Nezer and colleagues offer several reasonable suggestions for addressing the concerns that their work will raise.

The first is to share unthresholded activity maps, because this will allow image-based meta-analysis. The authors found that such an analysis, which aggregated information across teams, yielded consensus results, no doubt aided by the fact that the spatial patterns in the activity maps were highly correlated across groups.

Second is a call to publicly share both data and code, making it easier for others to attempt to reproduce a paper’s findings. In this regard, the authors model good behaviour by making all their data and processing pipelines publicly available.

Third is the use of pre-registration — in which a hypothesis and analysis plan is made public before the experiment is performed. It is unfortunately common for researchers to explore various pipelines to find the version that yields the ‘best’ results, ultimately reporting only that pipeline and ignoring the others. This practice can lead to errors and make it difficult to replicate findings4. Pre-registration would make it easy to detect cases in which researchers had explored various pipelines.

Fourth is to analyse all data through multiple pipelines and use the results to obtain consensus findings. This could be achieved by implementing the type of meta-analysis used by Botvinik-Nezer and co-workers.

Another goal of the study was to evaluate how accurately researchers could predict the number of teams that would report significant results for each hypothesis. To study this, the authors ran separate ‘prediction markets’, one for the analysis teams and one for researchers who did not participate in the analysis. In them, researchers attempted to predict the outcomes of the scientific analyses and received monetary payouts on the basis of how well they predicted performance. Participants — even researchers who had direct knowledge of the data set — consistently overestimated the likelihood of significant findings. Botvinik-Nezer et al. do not explicitly explore how the analysts’ prior beliefs affected their findings and pipeline choices. For example, if a research finding does not initially coincide with expectations, will groups seek to alter pipelines until expectations and results align? The prevalence of pipeline exploration implies that this is likely.

What other improvements could be made for the future? One approach is to use pipeline-optimization tools5,6 to reduce analysis flexibility. These tools automatically identify pipelines that maximize reproducibility, which can reduce the risk of excessive pipeline exploration and selective reporting. In addition, increased use of sensitivity analyses to evaluate the effects of pipeline decisions would provide a better understanding of the link between analysis choices and research findings. The fact that activity maps were highly correlated across groups implies that multivariate statistical approaches, which identify spatial patterns in the data, might provide more-consistent results across pipelines than would a series of tests performed at individual voxels.

Ultimately, neuroimaging results should be carefully verified using independent data sets to demonstrate generalizability across samples, research contexts and populations. A positive example of this already being done comes from an approach for developing predictive models on the basis of brain activation, which can be shared, tested in multiple contexts and used in applied settings7.

It seems unlikely that the fMRI field will ever coalesce on a standard workflow that is applicable to all types of study, because studies tend to be too varied for one pipeline to always be appropriate. But Botvinik-Nezer et al. conclude their paper by calling for an increased awareness of the situation, and a drive to improve the quality of method reporting. This is wise and prudent advice that researchers in any field analysing high-dimensional data would be well advised to heed.

doi: 10.1038/d41586-020-01282-z


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