The increasing interest in the investigation of social behaviours of a group of animals has heightened the need for developing tools that provide robust quantitative data. Drosophila melanogaster has emerged as an attractive model for behavioural analysis; however, there are still limited ways to monitor fly behaviour in a quantitative manner. To study social behaviour of a group of flies, acquiring the position of each individual over time is crucial. There are several studies that have tried to solve this problem and make this data acquisition automated. However, none of these studies has addressed the problem of keeping track of flies for a long period of time in three-dimensional space. Recently, we have developed an approach that enables us to detect and keep track of multiple flies in a three-dimensional arena for a long period of time, using multiple synchronized and calibrated cameras. After detecting flies in each view, correspondence between views is established using a novel approach we call the 'sequential Hungarian algorithm'. Subsequently, the three-dimensional positions of flies in space are reconstructed. We use the Hungarian algorithm and Kalman filter together for data association and tracking. We evaluated rigorously the system's performance for tracking and behaviour detection in multiple experiments, using from one to seven flies. Overall, this system presents a powerful new method for studying complex social interactions in a three-dimensional environment.
Across a diversity of animals, male seminal fluid coagulates upon ejaculation to form a hardened structure known as a copulatory plug. Previous studies suggest that copulatory plugs evolved as a mechanism for males to impede remating by females, but detailed investigations into the time course over which plugs survive in the female's reproductive tract are lacking. Here, we cross males from eight inbred strains to females from two inbred strains of house mice (Mus musculus domesticus). Plug survival was significantly affected by male genotype. Against intuition, plug survival time was negatively correlated with plug size: long-lasting plugs were small and relatively more susceptible to proteolysis. Plug size was associated with divergence in major protein composition of seminal vesicle fluid, suggesting that changes in gene expression may play an important role in plug dynamics. In contrast, we found no correlation to genetic variation in the protein-coding regions of five genes thought to be important in copulatory plug formation (Tgm4, Svs1, Svs2, Svs4 and Svs5). Our study demonstrates a complex relationship between copulatory plug characteristics and survival. We discuss several models to explain unexpected variation in plug phenotypes.
Although there is a heritable basis for many animal behaviors, the genetic architecture of behavioral variation in natural populations remains mostly unknown, particularly in vertebrates. We sought to identify the genetic basis for social affiliation in two populations of threespine sticklebacks (Gasterosteus aculeatus) that differ in their propensity to school. Marine sticklebacks from Japan school strongly whereas benthic sticklebacks from a lake in Canada are more solitary. Here, we expanded on our previous efforts to identify quantitative trait loci (QTL) for differences in schooling tendency. We tested fish multiple times in two assays that test different aspects of schooling tendency: 1) the model school assay, which presents fish with a school of eight model sticklebacks; and 2) the choice assay, in which fish are given a choice between the model school and a stationary artificial plant. We found low-to-moderate levels of repeatability, ranging from 0.1 to 0.5, in schooling phenotypes. To identify the genomic regions that contribute to differences in schooling tendency, we used QTL mapping in two types of crosses: benthic × marine backcrosses and an F2 intercross. We found two QTL for time spent with the school in the model school assay, and one QTL for number of approaches to the school in the choice assay. These QTL were on three different linkage groups, not previously linked to behavioral differences in sticklebacks. Our results highlight the importance of using multiple crosses and robust behavioral assays to uncover the genetic basis of behavioral variation in natural populations.
BackgroundIn vivo imaging and quantification of fluorescent reporter molecules is increasingly useful in biomedical research. For example, tracking animal movement in 3D with simultaneous quantification of fluorescent transgenic reporters allows for correlations between behavior, aging and gene expression. However implementation has been hindered in the past by the complexity of operating the systems.ResultsWe report significant technical improvements and user-friendly software (called FluoreScore) that enables tracking of 3D movement and the dynamics of gene expression in adult Drosophila, using two cameras and recorded GFP videos. Expression of a transgenic construct encoding eGFP was induced in free-moving adult flies using the Gene-Switch system and RU486 drug feeding. The time course of induction of eGFP expression was readily quantified from internal tissues including central nervous tissue.ConclusionsFluoreScore should facilitate a variety of future studies involving quantification of movement behaviors and fluorescent molecules in free-moving animals.
Sticklebacks have long been used as model organisms in behavioural biology. An important anti-predator behaviour in sticklebacks is schooling. We plan to use quantitative trait locus mapping to identify the genetic basis for differences in schooling behaviour between marine and benthic sticklebacks. To do this, we need to quantify the schooling behaviour of thousands of fish. We have developed a robust high-throughput video analysis method that allows us to screen a few thousand individuals automatically. We propose a non-local background modelling approach that allows us to detect and track sticklebacks and obtain the schooling parameters efficiently.
Senescence is associated with changes in gene expression, including the upregulation of stress response- and innate immune response-related genes. In addition, aging animals exhibit characteristic changes in movement behaviors including decreased gait speed and a deterioration in sleep/wake rhythms. Here, we describe methods for tracking Drosophila melanogaster movements in 3D with simultaneous quantification of fluorescent transgenic reporters. This approach allows for the assessment of correlations between behavior, aging, and gene expression as well as for the quantification of biomarkers of aging.
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