Intuitively most of us would agree that standardized conditions in scientific experiments ensure reproducibility. If I know how something was done and exactly replicate the setup and protocol, I should get the same results. The authors of a Perspective in the April issue of Nature Methods argue otherwise when it comes to behavioral experiments done with live animals.

Hanno Wϋrbel and his co-workers present their view that standardizing environmental conditions in animal experiments actually leads to poor reproducibility, since it makes the results more vulnerable to the most subtle local differences that can result in conflicting results between different labs. Instead, the authors advocate systematic environmental heterogenization. In their opinion this will increase reproducibility and thus decrease the ethical cost of animal experiments.

To prove their point the authors performed a data-mining experiment in which they group together the results from several behavioral tests performed in a multi-laboratory study with mice. They re-analyzed the published data by either maximizing or minimizing the environmental variation between the groups. The results from the heterogeneous group—those within the maximized environmental variation group—were less variable than those from the standardized group.

With this analysis Wϋrbel and co-workers challenge the current paradigm that standardization is essential in animal experiments. Their view is likely to spark discussion and we invite you to contribute to this discussion here.

It is always interesting to see how papers we publish are received by the community and one useful metric is the number of times a paper is viewed or downloaded. Authors also like to see this information when their paper was one of the most heavily viewed papers.

Each month we will report the original research papers we published that received the most unique views (HTML) and downloads (PDF) the previous month. This will be broken down between papers published in that month’s issue and papers published in prior months. Now for the most popular downloads for the month of February.

Top 4 research papers published in the February issue

1. Microfluidic control of cell pairing and fusion

2. Photoactivatable mCherry for high-resolution two-color fluorescence microscopy

3. A bright and photostable photoconvertible fluorescent protein

4. miRNA in situ hybridization in formaldehyde and EDC–fixed tissues

Top 10 research papers published prior to the February issue

1. Mapping and quantifying mammalian transcriptomes by RNA-Seq

2. Stable knockdown of microRNA in vivo by lentiviral vectors

3. Genome-wide analysis of transcription factor binding sites based on ChIP-Seq data

4. Stem cell transcriptome profiling via massive-scale mRNA sequencing

5. mirWIP: microRNA target prediction based on microRNA-containing ribonucleoprotein-enriched transcripts

6. Genome-wide profiles of STAT1 DNA association using chromatin immunoprecipitation and massively parallel sequencing

7. Whole-cell 3D STORM reveals interactions between cellular structures with nanometer-scale resolution

8. Bright far-red fluorescent protein for whole-body imaging

9. High-resolution mapping of copy-number alterations with massively parallel sequencing

10. A versatile tool for conditional gene expression and knockdown

It has been appreciated for a while that fruit flies interact with each other, but studying these interactions is quite difficult. In fact, this is a general problem in understanding animal behavior – the measurements are incredibly tedious to make and to quantify. In a paper published yesterday in Nature Methods (Dankert et al, 2009), Heiko Dankert, Pietro Perona, David Anderson and colleagues now automate the process, using the power of machine vision to analyse social interactions in pairs of fruit flies.

Put two wild-type male fruit flies together, and they will lunge, tussle and threaten each other, actions classified as aggressive (however, not all interactions between male flies are aggressive). Put a male and a female fruit fly together though and, predictably, their interactions will be largely of the courtship variety; the male circles the female, extends his wings in a courtship ‘song’, and sometimes, things work out!

Dankert et al show in their paper that they can take simple videos of fly-pairs, and use new software to accurately identify these stereotypical aggression and courtship actions. This allows them to observe many such fly-pairs and to describe these behaviors quantitatively. What’s more, the software detects expected differences in fly behavior. Male flies in which octopaminergic neurons have been silenced or where the fruitless gene is spliced into a female-specific form are much less aggressive. Male flies in which cholinergic neurons have been feminized now court other males. The software sees these differences.

Importantly, the measurements can be made quickly and without researcher bias or fatigue. For instance, Dankert et al estimate that experiments that would take them about a hundred hours to perform manually can now be completed in a few minutes. This opens up the study of these behaviors to genetic and other screens, and it will be interesting indeed to see how such studies play out.