An important concept in ethology is that complex behaviors can be constructed from a set of basic motor patterns. Identifying the set of patterns available to an animal is key to making quantitative descriptions of behavior that reflect the underlying motor system organization. We addressed these questions in zebrafish larvae, which swim in bouts that are naturally segmented in time. We developed a robust and general purpose clustering method (clusterdv) to ensure accurate identification of movement clusters and applied it to a dataset consisting of millions of swim bouts, captured at high temporal resolution from a comprehensive set of behavioral contexts. We identified a set of thirteen basic swimming patterns that are used flexibly in various combinations across different behavioral contexts and show that this classification can be used to dissect the sensorimotor transformations underlying larval social behavior and hunting. Furthermore, using the same approach at different levels in the behavioral hierarchy, we show that the set of swim bouts are themselves constructed from a basic set of tail movements and that bouts are executed in sequences specific to different behaviors.

Summary Vertebrate locomotion at different speeds is driven by descending excitatory connections to central pattern generators in the spinal cord. To investigate how these inputs determine locomotor kinematics, we used whole-field visual motion to drive zebrafish to swim at different speeds. Larvae match the stimulus speed by utilizing more locomotor events, or modifying kinematic parameters such as the duration and speed of swimming bouts, the tail-beat frequency, and choice of gait. We used laser ablations, electrical stimulation, and activity recordings in descending neurons of the nucleus of the medial longitudinal fasciculus (nMLF) to dissect their contribution to controlling forward movement. We found that the activity of single identified neurons within the nMLF is correlated with locomotor kinematics, and modulates both the duration and oscillation frequency of tail movements. By identifying the contribution of individual supraspinal circuit elements to locomotion kinematics we build a better understanding of how the brain controls movement.

Calcium imaging with cellular resolution typically requires an animal to be tethered under a microscope, which substantially restricts the range of behaviors that can be studied. To expand the behavioral repertoire amenable to imaging, we have developed a tracking microscope that enables whole-brain calcium imaging with cellular resolution in freely swimming larval zebrafish. This microscope uses infrared imaging to track a target animal in a behavior arena. On the basis of the predicted trajectory of the animal, we applied optimal control theory to a motorized stage system to cancel brain motion in three dimensions. We combined this motion-cancellation system with differential illumination focal filtering, a variant of HiLo microscopy, which enabled us to image the brain of a freely swimming larval zebrafish for more than an hour. This work expands the repertoire of natural behaviors that can be studied with cellular-resolution calcium imaging to potentially include spatial navigation, social behavior, feeding and reward.

Local Burden of Disease Child Growth Failure Collaborators* Childhood malnutrition is associated with high morbidity and mortality globally 1. Undernourished children are more likely to experience cognitive, physical, and metabolic developmental impairments that can lead to later cardiovascular disease, reduced intellectual ability and school attainment, and reduced economic productivity in adulthood 2. Child growth failure (CGF), expressed as stunting, wasting, and underweight in children under five years of age (0-59 months), is a specific subset of undernutrition characterized by insufficient height or weight against age-specific growth reference standards 3-5. The prevalence of stunting, wasting, or underweight in children under five is the proportion of children with a height-forage , weight-for-height, or weight-forage z-score, respectively, that is more than two standard deviations below the World Health Organization's median growth reference standards for a healthy population 6. Subnational estimates of CGF report substantial heterogeneity within countries, but are available primarily at the first administrative level (for example, states or provinces) 7 ; the uneven geographical distribution of CGF has motivated further calls for assessments that can match the local scale of many public health programmes 8. Building from our previous work mapping CGF in Africa 9 , here we provide the first, to our knowledge, mapped highspatial-resolution estimates of CGF indicators from 2000 to 2017 across 105 low-and middle-income countries (LMICs), where 99% of affected children live 1 , aggregated to policy-relevant first and second (for example, districts or counties) administrativelevel units and national levels. Despite remarkable declines over the study period, many LMICs remain far from the ambitious World Health Organization Global Nutrition Targets to reduce stunting by 40% and wasting to less than 5% by 2025. Large disparities in prevalence and progress exist across and within countries; our maps identify high-prevalence areas even within nations otherwise succeeding in reducing overall CGF prevalence. By highlighting where the highest-need populations reside, these geospatial estimates can support policy-makers in planning interventions that are adapted locally and in efficiently directing resources towards reducing CGF and its health implications. Despite improvements in nearly all LMICs, stunting remained the most widespread and prevalent indicator of CGF throughout the study period. Overall, estimated childhood stunting prevalence across LMICs decreased from 36.9% (95% uncertainty interval, 32.8-41.4%) in 2000 to 26.6% (21.5-32.4%) in 2017. Progress was particularly noticeable in Central America and the Caribbean, Andean South America, North Africa, and East Asia regions, and in some coastal central and western sub-Saharan African (SSA) countries, where most areas with estimated stunting prevalence of at least 50% in 2000 had reduced to 30% or less by 2017 (Fig. 1a, b). By 2017, zones with the highest prev...

SummaryAutoinducer-2 (AI-2) a signal produced by a range of phylogenetically distant microorganisms, enables inter-species cell-cell communication and regulates many bacterial phenotypes. Certain bacteria can interfere with AI-2-regulated behaviours of neighbouring species by internalizing AI-2 using the Lsr transport system (encoded by the lsr operon). AI-2 imported by the Lsr is phosphorylated by the LsrK kinase and AI-2-phosphate is the inducer of the lsr operon. Here we show that in Escherichia coli the phosphoenolpyruvate phosphotransferase system (PTS) is required for Lsr activation and is essential for AI-2 internalization. Although the phosphorylation state of Enzyme I of PTS is important for this regulation, LsrK is necessary for the phosphorylation of AI-2, indicating that AI-2 is not phosphorylated by PTS. Our results suggest that AI-2 internalization is initiated by a PTS-dependent mechanism, which provides sufficient intracellular AI-2 to relieve repression of the lsr operon and, thus induce depletion of AI-2 from the extracellular environment. The fact that AI-2 internalization is not only controlled by the communitydependent accumulation of AI-2, but also depends on the phosphorylation state of PTS suggests that E. coli can integrate information on the availability of substrates with external communal information to control quorum sensing and its interference.

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The molecule (S)-4,5-dihydroxy-2,3-pentanedione (DPD) is produced by many different species of bacteria and is the precursor of the signal molecule autoinducer-2 (AI-2). AI-2 mediates interspecies communication and facilitates regulation of bacterial behaviors such as biofilm formation and virulence. A variety of bacterial species have the ability to sequester and process the AI-2 present in their environment, thereby interfering with the cell-cell communication of other bacteria. This process involves the AI-2-regulated lsr operon, comprised of the Lsr transport system that facilitates uptake of the signal, a kinase that phosphorylates the signal to phospho-DPD (P-DPD), and enzymes (like LsrG) that are responsible for processing the phosphorylated signal. Because P-DPD is the intracellular inducer of the lsr operon, enzymes involved in P-DPD processing impact the levels of Lsr expression. Here we show that LsrG catalyzes isomerization of P-DPD into 3,4,4-trihydroxy-2-pentanone-5-phosphate. We present the crystal structure of LsrG, identify potential catalytic residues, and determine which of these residues affects P-DPD processing in vivo and in vitro. We also show that an lsrG deletion mutant accumulates at least 10 times more P-DPD than wild type cells. Consistent with this result, we find that the lsrG mutant has increased expression of the lsr operon and an altered profile of AI-2 accumulation and removal. Understanding of the biochemical mechanisms employed by bacteria to quench signaling of other species can be of great utility in the development of therapies to control bacterial behavior.Many bacteria regulate gene expression as a function of the density of the population. This process, called quorum sensing, enables these organisms to coordinate behaviors that are most beneficial when cells are working in unison (1, 2). Quorum sensing is mediated by signal molecules called autoinducers. One autoinducer (Autoinducer-2 (AI-2)3 ) is produced by many species of bacteria and can facilitate interspecies cell-cell signaling (3-5). AI-2 is produced by the enzyme LuxS, which synthesizes (S)-4,5-dihydroxy-2,3-pentanedione (DPD; Fig. 1A, 1), the linear form of a set of interconverting molecules with AI-2 activity (6 -8). AI-2 (or its synthase LuxS) has been shown to regulate important bacterial behaviors such as biofilm formation and the production of virulence factors (4, 5, 9, 10). Thus, one strategy for controlling these behaviors is to control the concentration (or availability) of AI-2. We have previously shown that certain bacteria can employ this strategy by processing AI-2 and thus quenching interspecies signaling (11). These bacteria include most members from the Enterobacteriaceae family (like the commensal Escherichia coli K12 and the pathogens E. coli O157 and Salmonella typhimurium) but also more distantly related bacteria such as the plant symbiont Sinorhizobium meliloti and the pathogen Bacillus anthracis (12).In these bacteria, AI-2 controls the expression of a system (named Lsr for luxS-regulated)...