Blue-green algae (Spirulina sp., Aphanizomenon flos-aquae) and Chlorella sp. are commercially distributed as organic algae dietary supplements. Cyanobacterial dietary products in particular have raised serious concerns, as they appeared to be contaminated with toxins e.g. microcystins (MCs) and consumers repeatedly reported adverse health effects following consumption of these products. The aim of this study was to determine the toxin contamination and the in vitro cytotoxicity of algae dietary supplement products marketed in Germany. In thirteen products consisting of Aph. flos-aquae, Spirulina and Chlorella or mixtures thereof, MCs, nodularins, saxitoxins, anatoxin-a and cylindrospermopsin were analyzed. Five products tested in an earlier market study were re-analyzed for comparison. Product samples were extracted and analyzed for cytotoxicity in A549 cells as well as for toxin levels by (1) phosphatase inhibition assay (PPIA), (2) Adda-ELISA and (3) LC-MS/MS. In addition, all samples were analyzed by PCR for the presence of the mcyE gene, a part of the microcystin and nodularin synthetase gene cluster. Only Aph. flos-aquae products were tested positive for MCs as well as the presence of mcyE. The contamination levels of the MC-positive samples were ≤ 1 μg MC-LR equivalents g(-1) dw. None of the other toxins were found in any of the products. However, extracts from all products were cytotoxic. In light of the findings, the distribution and commercial sale of Aph. flos-aquae products, whether pure or mixed formulations, for human consumption appear highly questionable.

The sensation of bitter substances can alert an animal that a specific type of food is harmful and should not be consumed. However, not all bitter compounds are equally toxic and some may even be beneficial in certain contexts. Thus, taste systems in general may have a broader range of functions than just in alerting the animal. In this study we investigate bitter sensing and processing in Drosophila larvae using quinine, a substance perceived by humans as bitter. We show that behavioral choice, feeding, survival, and associative olfactory learning are all directly affected by quinine. On the cellular level, we show that 12 gustatory sensory receptor neurons that express both GR66a and GR33a are required for quinine-dependent choice and feeding behavior. Interestingly, these neurons are not necessary for quinine-dependent survival or associative learning. On the molecular receptor gene level, the GR33a receptor, but not GR66a, is required for quinine-dependent choice behavior. A screen for gustatory sensory receptor neurons that trigger quinine-dependent choice behavior revealed that a single GR97a receptor gene expressing neuron located in the peripheral terminal sense organ is partially necessary and sufficient. For the first time, we show that the elementary chemosensory system of the Drosophila larva can serve as a simple model to understand the neuronal basis of taste information processing on the single cell level with respect to different behavioral outputs.

The Drosophila larva has a simple peripheral nervous system with a comparably small number of sensory neurons located externally at the head or internally along the pharynx to assess its chemical environment. It is assumed that larval taste coding occurs mainly via external organs (the dorsal, terminal, and ventral organ). However, the contribution of the internal pharyngeal sensory organs has not been explored. Here we find that larvae require a single pharyngeal gustatory receptor neuron pair called D1, which is located in the dorsal pharyngeal sensilla, in order to avoid caffeine and to associate an odor with caffeine punishment. In contrast, caffeine-driven reduction in feeding in non-choice situations does not require D1. Hence, this work provides data on taste coding via different receptor neurons, depending on the behavioral context. Furthermore, we show that the larval pharyngeal system is involved in bitter tasting. Using ectopic expressions, we show that the caffeine receptor in neuron D1 requires the function of at least four receptor genes: the putative co-receptors Gr33a, Gr66a, the putative caffeine-specific receptor Gr93a, and yet unknown additional molecular component(s). This suggests that larval taste perception is more complex than previously assumed already at the sensory level. Taste information from different sensory organs located outside at the head or inside along the pharynx of the larva is assembled to trigger taste guided behaviors.

In the last decade the Drosophila larva has evolved into a simple model organism offering the opportunity to integrate molecular genetics with systems neuroscience. This led to a detailed understanding of the neuronal networks for a number of sensory functions and behaviors including olfaction, vision, gustation and learning and memory. Typically, behavioral assays in use exploit simple Petri dish setups with either agarose or agar as a substrate. However, neither the quality nor the concentration of the substrate is generally standardized across these experiments and there is no data available on how larval behavior is affected by such different substrates. Here, we have investigated the effects of different agarose concentrations on several larval behaviors. We demonstrate that agarose concentration is an important parameter, which affects all behaviors tested: preference, feeding, learning and locomotion. Larvae can discriminate between different agarose concentrations, they feed differently on them, they can learn to associate an agarose concentration with an odor stimulus and change locomotion on a substrate of higher agarose concentration. Additionally, we have investigated the effect of agarose concentration on three quinine based behaviors: preference, feeding and learning. We show that in all cases examined the behavioral output changes in an agarose concentration-dependent manner. Our results suggest that comparisons between experiments performed on substrates differing in agarose concentration should be done with caution. It should be taken into consideration that the agarose concentration can affect the behavioral output and thereby the experimental outcomes per se potentially due to the initiation of an escape response or changes in foraging behavior on more rigid substrates.