The lumen of the small intestine (SI) is filled with particulates: microbes, therapeutic particles, and food granules. The structure of this particulate suspension could impact uptake of drugs and nutrients and the function of microorganisms; however, little is understood about how this suspension is re-structured as it transits the gut. Here, we demonstrate that particles spontaneously aggregate in SI luminal fluid ex vivo. We find that mucins and immunoglobulins are not required for aggregation. Instead, aggregation can be controlled using polymers from dietary fiber in a manner that is qualitatively consistent with polymer-induced depletion interactions, which do not require specific chemical interactions. Furthermore, we find that aggregation is tunable; by feeding mice dietary fibers of different molecular weights, we can control aggregation in SI luminal fluid. This work suggests that the molecular weight and concentration of dietary polymers play an underappreciated role in shaping the physicochemical environment of the gut.Editorial note: This article has been through an editorial process in which the authors decide how to respond to the issues raised during peer review. The Reviewing Editor's assessment is that all the issues have been addressed (see decision letter).

No abstract

17The lumen of the small intestine (SI) is filled with particulates: microbes, therapeutic particles, and food 18 granules. The structure of this particulate suspension could impact uptake of drugs and nutrients and the 19 function of microorganisms; however, little is understood about how this suspension is re-structured as it 20 transits the gut. Here, we demonstrate that particles spontaneously aggregate in SI luminal fluid ex vivo. We 21 find that mucins and immunoglobulins are not required for aggregation. Instead, aggregation can be controlled 22 using polymers from dietary fiber in a manner that is qualitatively consistent with polymer-induced depletion 23 interactions, which do not require specific chemical interactions. Furthermore, we find that aggregation is 24 tunable; by feeding mice dietary fibers of different molecular weights, we can control aggregation in SI luminal 25 fluid. This work suggests that the molecular weight and concentration of dietary polymers play an 26 underappreciated role in shaping the physicochemical environment of the gut. 27 28 Results 54PEG-coated particles aggregate in fluid from the murine small intestine 55 It has been observed that both bacteria (19)(20)(21)23,25,26) and particles (3,(36)(37)(38) aggregate in the gut. 56Experiments have been performed in which mice are orally co-administered carboxylate-coated nanoparticles, 57 which are mucoadhesive, and PEG-coated nanoparticles, which are mucus-penetrating (3). The carboxylate-58 coated particles formed large aggregates in the center of the gut lumen. In contrast, PEG-coated particles were 59 sometimes found co-localized with carboxylate-coated particles and also penetrated mucus, distributing across 60 the underlying epithelium of the SI as aggregates and single particles. 61To evaluate the distribution of particulate suspensions in the SI, we suspended 1-µm-diameter 62 fluorescent PEG-coated particles (see Materials and Methods for synthesis) in buffers isotonic to the SI and 63 orally administered them to mice. We chose 1 µm-diameter particles because of their similarity in size to 64 bacteria. We collected luminal contents after 3 h and confirmed using confocal fluorescence and reflectance 65 microscopy that these particles aggregated with each other and co-aggregated with what appeared to be digesta 66 (Fig. 1C and D; Materials and Methods). On separate mice, fluorescent scanning was used to verify that 67 particles do transit the SI after 3 h (Fig. 1A and B; Materials and Methods).68 69 109the sizes of all aggregates in solution using confocal microscopy (see Materials and Methods). From these 110 datasets, we created volume-weighted empirical cumulative distribution functions (ECDFs) of all the aggregate 111 sizes in a given solution. We used these volume-weighted ECDFs to compare the extent of aggregation in a 112 given sample ( Fig. 2F and H). To test the variability of aggregation in samples collected from groups of mice 113 treated under the same conditions, we compared the extent of aggregation in p...

Benzoquinones are a phylogenetically widespread compound class within arthropods, appearing in harvestman, millipedes and insects. Whereas the function of benzoquinones as defensive compounds against potential predators and microbes has been well established, the full extent of benzoquinone usage across arthropods, and especially within Insecta, has yet to be established. Adding to the growing list of unique evolutionary origins of benzoquinone employment, we describe in this paper the metathoracic scent gland secretion of the mirid bug Pamillia behrensii, which is composed of heptan-2-one, 2-heptyl acetate, 2,3-dimethyl-1-4-benzoquinone, 2,3-dimethyl-1-4-hydroquinone as well as one unknown compound. Similarly, to many other arthropods that use benzoquinones, Pamillia releases the contents of its gland as a defensive mechanism in response to harassment by other arthropod predators. Morphological investigation of the gland showed that the benzoquinone-producing gland complex of P. behrensii follows a similar blueprint to metathoracic scent glands described in other Heteropterans. Overall, our data further underpins the widespread convergent evolution and use of benzoquinones for defense across the Arthropoda, now including the order Hemiptera.

How evolution at the cellular level potentiates change at the macroevolutionary level is a major question in evolutionary biology. With >66,000 described species, rove beetles (Staphylinidae) comprise the largest metazoan family. Their exceptional radiation has been coupled to pervasive biosynthetic innovation whereby numerous lineages bear defensive glands with diverse chemistries. Here, we combine comparative genomic and single cell transcriptomic data from across the largest rove beetle clade, Aleocharinae. We retrace the functional evolution of two novel secretory cell types that together comprise the tergal gland - a putative catalyst behind Aleocharinae's megadiversity. We identify key genomic contingencies that were critical to the assembly of each cell type and their organ-level partnership in manufacturing the beetle's defensive secretion. This process hinged on evolving a mechanism for regulated production of noxious benzoquinones that appears convergent with plant toxin release systems, and synthesis of an effective benzoquinone solvent that weaponized the total secretion. We show that this cooperative biosynthetic system arose at the Jurassic-Cretaceous boundary, and that following its establishment, both cell types underwent ~150 million years of stasis, their chemistry and core molecular architecture maintained almost clade-wide as Aleocharinae radiated globally into tens of thousands of lineages. Despite this deep conservation, we show that the two cell types have acted as substrates for the emergence of adaptive, biochemical novelties - most dramatically in symbiotic lineages that have infiltrated social insect colonies and produce host behavior-manipulating secretions. Our findings uncover genomic and cell type evolutionary processes underlying the origin, functional conservation and evolvability of a chemical innovation in beetles.