Rewarding Capacity of Optogenetically Activating a Giant GABAergic Central-Brain Interneuron in Larval Drosophila.

Larvae of the fruit fly Drosophila melanogaster are a powerful study case for understanding the neural circuits underlying behavior. Indeed, the numerical simplicity of the larval brain has permitted the reconstruction of its synaptic connectome, and genetic tools for manipulating single, identified neurons allow neural circuit function to be investigated with relative ease and precision. We focus on one of the most complex neurons in the brain of the larva (of either sex), the GABAergic anterior paired lateral neuron (APL). Using behavioral and connectomic analyses, optogenetics, Ca2+ imaging, and pharmacology, we study how APL affects associative olfactory memory. We first provide a detailed account of the structure, regional polarity, connectivity, and metamorphic development of APL, and further confirm that optogenetic activation of APL has an inhibiting effect on its main targets, the mushroom body Kenyon cells. All these findings are consistent with the previously identified function of APL in the sparsening of sensory representations. To our surprise, however, we found that optogenetically activating APL can also have a strong rewarding effect. Specifically, APL activation together with odor presentation establishes an odor-specific, appetitive, associative short-term memory, whereas naive olfactory behavior remains unaffected. An acute, systemic inhibition of dopamine synthesis as well as an ablation of the dopaminergic pPAM neurons impair reward learning through APL activation. Our findings provide a study case of complex circuit function in a numerically simple brain, and suggest a previously unrecognized capacity of central-brain GABAergic neurons to engage in dopaminergic reinforcement.SIGNIFICANCE STATEMENT The single, identified giant anterior paired lateral (APL) neuron is one of the most complex neurons in the insect brain. It is GABAergic and contributes to the sparsening of neuronal activity in the mushroom body, the memory center of insects. We provide the most detailed account yet of the structure of APL in larval Drosophila as a neurogenetically accessible study case. We further reveal that, contrary to expectations, the experimental activation of APL can exert a rewarding effect, likely via dopaminergic reward pathways. The present study both provides an example of unexpected circuit complexity in a numerically simple brain, and reports an unexpected effect of activity in central-brain GABAergic circuits.

IR76b-expressing neurons in Drosophila melanogaster are necessary for associative reward learning of an amino acid mixture.

Learning where to find nutrients while at the same time avoiding toxic food is essential for survival of any animal. Using Drosophila melanogaster larvae as a study case, we investigate the role of gustatory sensory neurons expressing IR76b for associative learning of amino acids, the building blocks of proteins. We found surprising complexity in the neuronal underpinnings of sensing amino acids, and a functional division of sensory neurons. We found that the IR76b receptor is dispensable for amino acid learning, whereas the neurons expressing IR76b are specifically required for the rewarding but not the punishing effect of amino acids. This unexpected dissociation in neuronal processing of amino acids for different behavioural functions provides a study case for functional divisions of labour in gustatory systems.

Testing for concordance between predicted species richness, past prioritization, and marine protected area designations in the western Indian Ocean.

Scientific advances in environmental data coverage and machine learning algorithms have improved the ability to make large-scale predictions where data are missing. These advances allowed us to develop a spatially resolved proxy for predicting numbers of tropical nearshore marine taxa. A diverse marine environmental spatial database was used to model numbers of taxa from ∼1000 field sites, and the predictions were applied to all 7039 6.25-km2 reef cells in 9 ecoregions and 11 nations of the western Indian Ocean. Our proxy for total numbers of taxa was based on the positive correlation (r2 = 0.24) of numbers of taxa of hard corals and 5 highly diverse reef fish families. Environmental relationships indicated that the number of fish species was largely influenced by biomass, nearness to people, governance, connectivity, and productivity and that coral taxa were influenced mostly by physicochemical environmental variability. At spatial delineations of province, ecoregion, nation, and strength of spatial clustering, we compared areas of conservation priority based on our total species proxy with those identified in 3 previous priority-setting reports and with the protected area database. Our method identified 119 locations that fit 3 numbers of taxa (hard coral, fish, and their combination) and 4 spatial delineations (nation, ecoregion, province, and reef clustering) criteria. Previous publications on priority setting identified 91 priority locations of which 6 were identified by all reports. We identified 12 locations that fit our 12 criteria and corresponded with 3 previously identified locations, 65 that aligned with at least 1 past report, and 28 that were new locations. Only 34% of the 208 marine protected areas in this province overlapped with identified locations with high numbers of predicted taxa. Differences occurred because past priorities were frequently based on unquantified perceptions of remoteness and preselected priority taxa. Our environment-species proxy and modeling approach can be considered among other important criteria for making conservation decisions.

Evaluación de la concordancia entre la riqueza de especies pronosticada, priorizaciones pasadas y la designación de áreas marinas protegidas en el oeste del Océano Índico Resumen Los avances científicos en la cobertura de datos ambientales y los algoritmos de aprendizaje automatizado han mejorado la capacidad de predecir a gran escala cuando hacen falta datos. Estos avances nos permiten desarrollar un representante con resolución espacial para predecir la cantidad de taxones marinos en las costas tropicales. Usamos una base de datos espaciales de diversos ambientes marinos para modelar la cantidad de taxones a partir de ∼1000 sitios de campo y aplicamos las predicciones a las 7039 celdas arrecifales de 6.25­km2 en nueve ecorregiones y once países del oeste del Océano Índico. Nuestro representante para la cantidad total de taxones se basó en la correlación positiva (r2=0.24) de la cantidad de taxones de corales duros y cinco familias de peces arrecifales con diversidad alta. Las relaciones ambientales indicaron que el número de especies de peces estuvo influenciado principalmente por la biomasa, la cercanía a las personas, la gestión, la conectividad y la productividad y que los taxones de coral estuvieron influenciados principalmente por la variabilidad ambiental fisicoquímica. Comparamos la prioridad de las áreas de conservación a nivel de las delimitaciones espaciales de provincia, ecorregión, nación y fuerza del agrupamiento espacial basado en nuestro total de especies representantes con aquellas especies identificadas en tres reportes previos de establecimiento de prioridades y con la base de datos de áreas protegidas. Con nuestro método identificamos 119 localidades aptas para tres cantidades de taxones (corales duros, peces y su combinación) y cuatro criterios de delimitación espacial (nación, ecorregión, provincia y grupo de arrecifes). Las publicaciones previas sobre el establecimiento de prioridades identificaron 91 localidades prioritarias de las cuales seis fueron identificadas por todos los reportes. Identificamos doce localidades que se ajustan a nuestros doce criterios y se correspondieron con tres localidades identificadas previamente, 65 que se alinearon con al menos un reporte anterior y 28 que eran nuevas localidades. Sólo 34% de las 208 áreas marinas protegidas en esta provincia se traslaparon con localidades identificadas con un gran número de taxones pronosticados. Hubo diferencias porque en el pasado se priorizaba frecuentemente con base en las percepciones no cuantificadas de lo remoto y prioritario de los taxones preseleccionados. Nuestra especie representante del ambiente y nuestra estrategia de modelo pueden considerarse entre otros criterios importantes para tomar decisiones de conservación.

Optogenetically induced reward and 'frustration' memory in larval Drosophila melanogaster.

Animals, including humans, form oppositely valenced memories for stimuli that predict the occurrence versus the termination of a reward: appetitive 'reward' memory for stimuli associated with the occurrence of a reward and aversive 'frustration' memory for stimuli that are associated with its termination. We characterized these memories in larval Drosophila melanogaster using a combination of Pavlovian conditioning, optogenetic activation of the dopaminergic central-brain DAN-i1864 neuron, and high-resolution video-tracking. This reveals their dependency on the number of training trials and the duration of DAN-i1864 activation, their temporal stability, and the parameters of locomotion that are modulated during memory expression. Together with previous results on 'punishment' versus 'relief' learning by DAN-f1 neuron activation, this reveals a 2×2 matrix of timing-dependent memory valence for the occurrence/termination of reward/punishment. These findings should aid the understanding and modelling of how brains decipher the predictive, causal structure of events around a target reinforcing occurrence.

Identification of Dopaminergic Neurons That Can Both Establish Associative Memory and Acutely Terminate Its Behavioral Expression.

An adaptive transition from exploring the environment in search of vital resources to exploiting these resources once the search was successful is important to all animals. Here we study the neuronal circuitry that allows larval Drosophila melanogaster of either sex to negotiate this exploration-exploitation transition. We do so by combining Pavlovian conditioning with high-resolution behavioral tracking, optogenetic manipulation of individually identified neurons, and EM data-based analyses of synaptic organization. We find that optogenetic activation of the dopaminergic neuron DAN-i1 can both establish memory during training and acutely terminate learned search behavior in a subsequent recall test. Its activation leaves innate behavior unaffected, however. Specifically, DAN-i1 activation can establish associative memories of opposite valence after paired and unpaired training with odor, and its activation during the recall test can terminate the search behavior resulting from either of these memories. Our results further suggest that in its behavioral significance DAN-i1 activation resembles, but does not equal, sugar reward. Dendrogram analyses of all the synaptic connections between DAN-i1 and its two main targets, the Kenyon cells and the mushroom body output neuron MBON-i1, further suggest that the DAN-i1 signals during training and during the recall test could be delivered to the Kenyon cells and to MBON-i1, respectively, within previously unrecognized, locally confined branching structures. This would provide an elegant circuit motif to terminate search on its successful completion.SIGNIFICANCE STATEMENT In the struggle for survival, animals have to explore their environment in search of food. Once food is found, however, it is adaptive to prioritize exploiting it over continuing a search that would now be as pointless as searching for the glasses you are wearing. This exploration-exploitation trade-off is important for animals and humans, as well as for technical search devices. We investigate which of the only 10,000 neurons of a fruit fly larva can tip the balance in this trade-off, and identify a single dopamine neuron called DAN-i1 that can do so. Given the similarities in dopamine neuron function across the animal kingdom, this may reflect a general principle of how search is terminated once it is successful.

One-trial learning in larval Drosophila.

Animals of many species are capable of "small data" learning, that is, of learning without repetition. Here we introduce larval Drosophila melanogaster as a relatively simple study case for such one-trial learning. Using odor-food associative conditioning, we first show that a sugar that is both sweet and nutritious (fructose) and sugars that are only sweet (arabinose) or only nutritious (sorbitol) all support appetitive one-trial learning. The same is the case for the optogenetic activation of a subset of dopaminergic neurons innervating the mushroom body, the memory center of the insects. In contrast, no one-trial learning is observed for an amino acid reward (aspartic acid). As regards the aversive domain, one-trial learning is demonstrated for high-concentration sodium chloride, but is not observed for a bitter tastant (quinine). Second, we provide follow-up, parametric analyses of odor-fructose learning. Specifically, we ascertain its dependency on the number and duration of training trials, the requirements for the behavioral expression of one-trial odor-fructose memory, its temporal stability, and the feasibility of one-trial differential conditioning. Our results set the stage for a neurogenetic analysis of one-trial learning and define the requirements for modeling mnemonic processes in the larva.

Reversal learning in Drosophila larvae.

Adjusting behavior to changed environmental contingencies is critical for survival, and reversal learning provides an experimental handle on such cognitive flexibility. Here, we investigate reversal learning in larval Drosophila Using odor-taste associations, we establish olfactory reversal learning in the appetitive and the aversive domain, using either fructose as a reward or high-concentration sodium chloride as a punishment, respectively. Reversal learning is demonstrated both in differential and in absolute conditioning, in either valence domain. In differential conditioning, the animals are first trained such that an odor A is paired, for example, with the reward whereas odor B is not (A+/B); this is followed by a second training phase with reversed contingencies (A/B+). In absolute conditioning, odor B is omitted, such that the animals are first trained with paired presentations of A and reward, followed by unpaired training in the second training phase. Our results reveal "true" reversal learning in that the opposite associative effects of both the first and the second training phase are detectable after reversed-contingency training. In what is a surprisingly quick, one-trial contingency adjustment in the Drosophila larva, the present study establishes a simple and genetically easy accessible study case of cognitive flexibility.

Drivers of Soft and Stony Coral Community Distribution on the High-Latitude Coral Reefs of South Africa.

The role of abiotic parameters in determining the distribution of coral communities was assessed on the relatively pristine Maputaland reefs of South Africa from comprehensive reef survey data. The reefs, on which 42 communities could be defined, occur within three geographically separate complexes. Patterns in benthic distribution could be partially explained by latitude and depth, in particular, with slope, turbulence and reef aspect playing far less role in that order. A few species were associated exclusively or in high abundance with some of the communities; in most, it was the ratio of otherwise cosmopolitan species within all of the reef complexes that distinguished them. Complex biotic variables were also considered but not quantified and it is likely that low genetic connectivity and high levels of self-seeding result in a measure of isolation of the communities within a latitudinal gradient.

Common microbehavioral "footprint" of two distinct classes of conditioned aversion.

Avoiding unfavorable situations is a vital skill and a constant task for any animal. Situations can be unfavorable because they feature something that the animal wants to escape from, or because they do not feature something that it seeks to obtain. We investigate whether the microbehavioral mechanisms by which these two classes of aversion come about are shared or distinct. We find that larval Drosophila avoid odors either previously associated with a punishment, or previously associated with the lack of a reward. These two classes of conditioned aversion are found to be strikingly alike at the microbehavioral level. In both cases larvae show more head casts when oriented toward the odor source than when oriented away, and direct fewer of their head casts toward the odor than away when oriented obliquely to it. Thus, conditioned aversion serving two qualitatively different functions-escape from a punishment or search for a reward-is implemented by the modulation of the same microbehavioral features. These features also underlie conditioned approach, albeit with opposite sign. That is, the larvae show conditioned approach toward odors previously associated with a reward, or with the lack of a punishment. In order to accomplish both these classes of conditioned approach the larvae show fewer head casts when oriented toward an odor, and direct more of their head casts toward it when they are headed obliquely. Given that the Drosophila larva is a genetically tractable model organism that is well suited to study simple circuits at the single-cell level, these analyses can guide future research into the neuronal circuits underlying conditioned approach and aversion, and the computational principles of conditioned search and escape.

The Ol1mpiad: concordance of behavioural faculties of stage 1 and stage 3 Drosophila larvae.

Mapping brain function to brain structure is a fundamental task for neuroscience. For such an endeavour, the Drosophila larva is simple enough to be tractable, yet complex enough to be interesting. It features about 10,000 neurons and is capable of various taxes, kineses and Pavlovian conditioning. All its neurons are currently being mapped into a light-microscopical atlas, and Gal4 strains are being generated to experimentally access neurons one at a time. In addition, an electron microscopic reconstruction of its nervous system seems within reach. Notably, this electron microscope-based connectome is being drafted for a stage 1 larva - because stage 1 larvae are much smaller than stage 3 larvae. However, most behaviour analyses have been performed for stage 3 larvae because their larger size makes them easier to handle and observe. It is therefore warranted to either redo the electron microscopic reconstruction for a stage 3 larva or to survey the behavioural faculties of stage 1 larvae. We provide the latter. In a community-based approach we called the Ol1mpiad, we probed stage 1 Drosophila larvae for free locomotion, feeding, responsiveness to substrate vibration, gentle and nociceptive touch, burrowing, olfactory preference and thermotaxis, light avoidance, gustatory choice of various tastants plus odour-taste associative learning, as well as light/dark-electric shock associative learning. Quantitatively, stage 1 larvae show lower scores in most tasks, arguably because of their smaller size and lower speed. Qualitatively, however, stage 1 larvae perform strikingly similar to stage 3 larvae in almost all cases. These results bolster confidence in mapping brain structure and behaviour across developmental stages.

Synapsin is required to "boost" memory strength for highly salient events.

Synapsin is an evolutionarily conserved presynaptic phosphoprotein. It is encoded by only one gene in the Drosophila genome and is expressed throughout the nervous system. It regulates the balance between reserve and releasable vesicles, is required to maintain transmission upon heavy demand, and is essential for proper memory function at the behavioral level. Task-relevant sensorimotor functions, however, remain intact in the absence of Synapsin. Using an odor-sugar reward associative learning paradigm in larval Drosophila, we show that memory scores in mutants lacking Synapsin (syn(97)) are lower than in wild-type animals only when more salient, higher concentrations of odor or of the sugar reward are used. Furthermore, we show that Synapsin is selectively required for larval short-term memory. Thus, without Synapsin Drosophila larvae can learn and remember, but Synapsin is required to form memories that match in strength to event salience-in particular to a high saliency of odors, of rewards, or the salient recency of an event. We further show that the residual memory scores upon a lack of Synapsin are not further decreased by an additional lack of the Sap47 protein. In combination with mass spectrometry data showing an up-regulated phosphorylation of Synapsin in the larval nervous system upon a lack of Sap47, this is suggestive of a functional interdependence of Synapsin and Sap47.

The impact of odor-reward memory on chemotaxis in larval Drosophila.

How do animals adaptively integrate innate with learned behavioral tendencies? We tackle this question using chemotaxis as a paradigm. Chemotaxis in the Drosophila larva largely results from a sequence of runs and oriented turns. Thus, the larvae minimally need to determine (i) how fast to run, (ii) when to initiate a turn, and (iii) where to direct a turn. We first report how odor-source intensities modulate these decisions to bring about higher levels of chemotactic performance for higher odor-source intensities during innate chemotaxis. We then examine whether the same modulations are responsible for alterations of chemotactic performance by learned odor "valence" (understood throughout as level of attractiveness). We find that run speed (i) is neither modulated by the innate nor by the learned valence of an odor. Turn rate (ii), however, is modulated by both: the higher the innate or learned valence of the odor, the less often larvae turn whenever heading toward the odor source, and the more often they turn when heading away. Likewise, turning direction (iii) is modulated concordantly by innate and learned valence: turning is biased more strongly toward the odor source when either innate or learned valence is high. Using numerical simulations, we show that a modulation of both turn rate and of turning direction is sufficient to account for the empirically found differences in preference scores across experimental conditions. Our results suggest that innate and learned valence organize adaptive olfactory search behavior by their summed effects on turn rate and turning direction, but not on run speed. This work should aid studies into the neural mechanisms by which memory impacts specific aspects of behavior.

Identification of a bacterial pathogen associated with Porites white patch syndrome in the Western Indian Ocean.

Porites white patch syndrome (PWPS) is a coral disease recently described in the Western Indian Ocean. This study aimed to isolate and identify potential pathogens associated with PWPS utilizing both culture and nonculture screening techniques and inoculation trials. A total of 14 bacterial strains (those dominant in disease lesions, absent or rare in healthy tissues and considered potential pathogens in a previous study) were cultured and used to experimentally inoculate otherwise healthy individuals in an attempt to fulfil Henle-Koch's postulates. However, only one (P180R), identified as closely related (99-100% sequence identity based on 1.4 kb 16S RNA sequence) to Vibrio tubiashii, elicited signs of disease in tank experiments. Following experimental infection (which resulted in a 90% infection rate), the pathogen was also successfully re-isolated from the diseased tissues and re-inoculated in healthy corals colonies, therefore fulfilling the final stages of Henle-Koch's postulates. Finally, we report that PWPS appears to be a temperature-dependent disease, with significantly higher tissue loss (anova: d.f. = 2, F = 39.77, P < 0.01) occurring at 30 °C [1.45 ± 0.85 cm(2) per day (mean ± SE)] compared to ambient temperatures of 28 and 26 °C (0.73 ± 0.80 cm(2) per day (mean ± SE) and 0.51 ± 0.50 cm(2) per day (mean ± SE), respectively).

Identification and prevalence of coral diseases on three Western Indian Ocean coral reefs.

Coral diseases have caused a substantial decline in the biodiversity and abundance of reef-building corals. To date, more than 30 distinct diseases of scleractinian corals have been reported, which cause progressive tissue loss and/or affect coral growth, reproductive capacity, recruitment, species diversity and the abundance of reef-associated organisms. While coral disease research has increased over the last 4 decades, very little is known about coral diseases in the Western Indian Ocean. Surveys conducted at multiple sites in Reunion, South Africa and Mayotte between August 2010 and June 2012 revealed the presence of 6 main coral diseases: black band disease (BBD), white syndrome (WS), pink line syndrome (PLS), growth anomalies (GA), skeleton eroding band (SEB) and Porites white patch syndrome (PWPS). Overall, disease prevalence was higher in Reunion (7.5 ± 2.2%; mean ± SE) compared to South Africa (3.9 ± 0.8%) and Mayotte (2.7 ± 0.3%). Across locations, Acropora and Porites were the genera most susceptible to disease. Spatial variability was detected in both Reunion and South Africa, with BBD and WS more prevalent on shallow than deep reefs. There was also evidence of seasonality in 2 diseases: the prevalence of BBD and WS was higher in summer than winter. This was the first study to investigate the ecology of coral diseases, providing both qualitative and quantitative data, on Western Indian Ocean reefs, and surveys should be expanded to confirm these patterns.

Bitter-sweet processing in larval Drosophila.

"Sweet-" and "bitter-" tasting substances distinctively support attractive and aversive choice behavior, respectively, and therefore are thought to be processed by distinct pathways. Interestingly, electrophysiological recordings in adult Drosophila suggest that bitter and salty tastants, in addition to activating bitter, salt, or bitter/salt sensory neurons, can also inhibit sweet-sensory neurons. However, the behavioral significance of such a potential for combinatorial coding is little understood. Using larval Drosophila as a study case, we find that the preference towards fructose is inhibited when assayed in the background of the bitter tastant quinine. When testing the influence of quinine on the preference to other, equally preferred sweet tastants, we find that these sweet tastants differ in their susceptibility to be inhibited by quinine. Such stimulus specificity argues that the inhibitory effect of quinine is not due to general effects on locomotion or nausea. In turn, not all bitter tastants have the same potency to inhibit sweet preference; notably, their inhibitory potency is not determined by the strength of the avoidance of them. Likewise, equally avoided concentrations of sodium chloride differ in their potency to inhibit sugar preference. Furthermore, Gr33a-Gal4-positive neurons, while being necessary for bitter avoidance, are dispensable for inhibition of the sweet pathway. Thus, interactions across taste modalities are behaviorally significant and, as we discuss, arguably diverse in mechanism. These results suggest that the coding of tastants and the organization of gustatory behavior may be more combinatorial than is generally acknowledged.

Prediction error drives associative learning and conditioned behavior in a spiking model of Drosophila larva.

Predicting reinforcement from sensory cues is beneficial for goal-directed behavior. In insect brains, underlying associations between cues and reinforcement, encoded by dopaminergic neurons, are formed in the mushroom body. We propose a spiking model of the Drosophila larva mushroom body. It includes a feedback motif conveying learned reinforcement expectation to dopaminergic neurons, which can compute prediction error as the difference between expected and present reinforcement. We demonstrate that this can serve as a driving force in learning. When combined with synaptic homeostasis, our model accounts for theoretically derived features of acquisition and loss of associations that depend on the intensity of the reinforcement and its temporal proximity to the cue. From modeling olfactory learning over the time course of behavioral experiments and simulating the locomotion of individual larvae toward or away from odor sources in a virtual environment, we conclude that learning driven by prediction errors can explain larval behavior.

Maggot learning and Synapsin function.

Drosophila larvae are focused on feeding and have few neurons. Within these bounds, however, there still are behavioural degrees of freedom. This review is devoted to what these elements of flexibility are, and how they come about. Regarding odour-food associative learning, the emerging working hypothesis is that when a mushroom body neuron is activated as a part of an odour-specific set of mushroom body neurons, and coincidently receives a reinforcement signal carried by aminergic neurons, the AC-cAMP-PKA cascade is triggered. One substrate of this cascade is Synapsin, and therefore this review features a general and comparative discussion of Synapsin function. Phosphorylation of Synapsin ensures an alteration of synaptic strength between this mushroom body neuron and its target neuron(s). If the trained odour is encountered again, the pattern of mushroom body neurons coding this odour is activated, such that their modified output now allows conditioned behaviour. However, such an activated memory trace does not automatically cause conditioned behaviour. Rather, in a process that remains off-line from behaviour, the larvae compare the value of the testing situation (based on gustatory input) with the value of the odour-activated memory trace (based on mushroom body output). The circuit towards appetitive conditioned behaviour is closed only if the memory trace suggests that tracking down the learned odour will lead to a place better than the current one. It is this expectation of a positive outcome that is the immediate cause of appetitive conditioned behaviour. Such conditioned search for reward corresponds to a view of aversive conditioned behaviour as conditioned escape from punishment, which is enabled only if there is something to escape from - much in the same way as we only search for things that are not there, and run for the emergency exit only when there is an emergency. One may now ask whether beyond 'value' additional information about reinforcement is contained in the memory trace, such as information about the kind and intensity of the reinforcer used. The Drosophila larva may allow us to develop satisfyingly detailed accounts of such mnemonic richness - if it exists.

High-resolution analysis of individual Drosophila melanogaster larvae uncovers individual variability in locomotion and its neurogenetic modulation.

Neuronally orchestrated muscular movement and locomotion are defining faculties of multicellular animals. Due to its simple brain and genetic accessibility, the larva of the fruit fly Drosophila melanogaster allows one to study these processes at tractable levels of complexity. However, although the faculty of locomotion clearly pertains to the individual, most studies of locomotion in larvae use measurements aggregated across animals, or animals tested one by one, an extravagance for larger-scale analyses. This prevents grasping the inter- and intra-individual variability in locomotion and its neurogenetic determinants. Here, we present the IMBA (individual maggot behaviour analyser) for analysing the behaviour of individual larvae within groups, reliably resolving individual identity across collisions. We use the IMBA to systematically describe the inter- and intra-individual variability in locomotion of wild-type animals, and how the variability is reduced by associative learning. We then report a novel locomotion phenotype of an adhesion GPCR mutant. We further investigated the modulation of locomotion across repeated activations of dopamine neurons in individual animals, and the transient backward locomotion induced by brief optogenetic activation of the brain-descending 'mooncrawler' neurons. In summary, the IMBA is an easy-to-use toolbox allowing an unprecedentedly rich view of the behaviour and its variability of individual larvae, with utility in multiple biomedical research contexts.

A behavior-based circuit model of how outcome expectations organize learned behavior in larval Drosophila.

Drosophila larvae combine a numerically simple brain, a correspondingly moderate behavioral complexity, and the availability of a rich toolbox for transgenic manipulation. This makes them attractive as a study case when trying to achieve a circuit-level understanding of behavior organization. From a series of behavioral experiments, we suggest a circuitry of chemosensory processing, odor-tastant memory trace formation, and the "decision" process to behaviorally express these memory traces--or not. The model incorporates statements about the neuronal organization of innate vs. conditioned chemosensory behavior, and the types of interaction between olfactory and gustatory pathways during the establishment as well as the behavioral expression of odor-tastant memory traces. It in particular suggests that innate olfactory behavior is responsive in nature, whereas conditioned olfactory behavior is captured better when seen as an action in pursuit of its outcome. It incorporates the available neuroanatomical and behavioral data and thus should be useful as scaffold for the ongoing investigations of the chemo-behavioral system in larval Drosophila.

Accumulation of commonly used agricultural herbicides in coral reef organisms from iSimangaliso Wetland Park, South Africa.

Coral reefs are amongst the most biodiverse ecosystems on earth, but are significantly impacted by agricultural runoff. Despite herbicides being commonly detected in coastal waters, the possibility of herbicide accumulation in coral reef species has largely been overlooked. We investigate the accumulation of several herbicides in five species of coral reef invertebrates collected from ten sites along the Maputaland coast, South Africa. Multiple herbicide residues were detected in 95% of the samples, with total average concentrations across sites ranging between 25.2 ng g-1 to 51.3 ng g-1 dw. Acetochlor, alachlor and hexazinone were the predominant herbicides detected at all sites, with atrazine and simazine detected less frequently. Significant interactive effects were detected between sites nested in reef complex crossed with species, based on multiple and total herbicide concentrations. In general, multivariate herbicide concentrations varied significantly between species within and across most sites. Contrastingly, the concentrations of the different herbicides and that of total herbicide did not differ between conspecifics at most sites nested in their respective reef complexes. On average, highest total herbicide concentrations were measured in soft coral (Sarcophyton glaucum; 90.4 ± 60 ng g-1 and Sinularia gravis; 42.7 ± 25 ng g-1) and sponge (Theonela swinhoei; 39.0 ± 40 ng g-1) species, while significantly lower concentrations were detected in hard corals (Echinopora hirsutissima; 10.5 ± 5.9 ng g-1 and Acropora austera; 5.20 ± 4.5 ng g-1) at most sites. Agricultural runoff entering the ocean via the uMfolozi-St Lucia Estuary and Maputo Bay are likely sources of herbicide contamination to coral reefs in the region. There is an urgent need to assess the long-term effects of herbicide exposure on coral reef communities.