A single mutation weakens symbiont-induced reproductive manipulation through reductions in deubiquitylation efficiency

Animals interact with microbes that affect their performance and fitness, including endosymbionts that reside inside their cells. Maternally transmitted Wolbachia bacteria are the most common known endosymbionts, in large part because of their manipulation of host reproduction. For example, many Wolbachia cause cytoplasmic incompatibility (CI) that reduces host embryonic viability when Wolbachia-modified sperm fertilize uninfected eggs. Operons termed cifs control CI, and a single factor (cifA) rescues it, providing Wolbachia-infected females a fitness advantage. Despite CI’s prevalence in nature, theory indicates that natural selection does not act to maintain CI, which varies widely in strength. Here, we investigate the genetic and functional basis of CI-strength variation observed among sister Wolbachia that infect Drosophila melanogaster subgroup hosts. We cloned, Sanger sequenced, and expressed cif repertoires from weak CI–causing wYak in Drosophila yakuba, revealing mutations suspected to weaken CI relative to model wMel in D. melanogaster. A single valine-to-leucine mutation within the deubiquitylating (DUB) domain of the wYak cifB homolog (cidB) ablates a CI-like phenotype in yeast. The same mutation reduces both DUB efficiency in vitro and transgenic CI strength in the fly, each by about twofold. Our results map hypomorphic transgenic CI to reduced DUB activity and indicate that deubiquitylation is central to CI induction in cid systems. We also characterize effects of other genetic variation distinguishing wMel-like cifs. Importantly, CI strength determines Wolbachia prevalence in natural systems and directly influences the efficacy of Wolbachia biocontrol strategies in transinfected mosquito systems. These approaches rely on strong CI to reduce human disease.

Many endosymbionts spread through host populations by manipulating their reproduction. For example, Rickettsiella (1), Mesenet (2), Cardinium (3), and Wolbachia (4) all cause cytoplasmic incompatibility (CI) that reduces the viability of uninfected host embryos fertilized by symbiont-modified sperm (5–8). CI is common among Wolbachia bacterial strains, being observed in at least 10 arthropod orders (6). CI strength influences Wolbachia prevalence, with stronger CI producing higher Wolbachia infection frequencies in host populations (8, 9). Indeed, CI contributes significantly to Wolbachia’s status as the most-common known endosymbionts in nature (10).CI strength directly influences the efficacy of Wolbachia biocontrol programs, with vector-control groups relying on strong CI to either suppress mosquito populations (11, 12) or to transform them with pathogen-blocking Wolbachia like wMel that naturally infects Drosophila melanogaster (13–15). The World Health Organization recommends further developing these programs (16), which are currently protecting seven million people from disease with a goal of protecting half a billion by 2030 (14, 17).Operons generally termed cifs control CI (cifA/B) (5, 18–22), and CI induction can be rescued by one factor (cifA) (5, 19, 23). Theory indicates that natural selection does not act to increase or maintain CI (24), which varies considerably among even very closely related Wolbachia (25–27), potentially due to mutational erosion of cifs (28). For example, CI strength differs significantly among model wMel from Drosophila melanogaster and closely related wMel-like Wolbachia in the Drosophila yakuba clade (wYak, wSan, and wTei) that wMel diverged from in only the last 30,000 y (25, 27, 29, 30). We sought to determine how much and why naturally observed mutations in wMel-like cifs influence CI strength.     

Native microbiome impedes vertical transmission of Wolbachia in Anopheles mosquitoes

Over evolutionary time, Wolbachia has been repeatedly transferred between host species contributing to the widespread distribution of the symbiont in arthropods. For novel infections to be maintained, Wolbachia must infect the female germ line after being acquired by horizontal transfer. Although mechanistic examples of horizontal transfer exist, there is a poor understanding of factors that lead to successful vertical maintenance of the acquired infection. Using Anopheles mosquitoes (which are naturally uninfected by Wolbachia) we demonstrate that the native mosquito microbiota is a major barrier to vertical transmission of a horizontally acquired Wolbachia infection. After injection into adult Anopheles gambiae, some strains of Wolbachia invade the germ line, but are poorly transmitted to the next generation. In Anopheles stephensi, Wolbachia infection elicited massive blood meal-induced mortality, preventing development of progeny. Manipulation of the mosquito microbiota by antibiotic treatment resulted in perfect maternal transmission at significantly elevated titers of the wAlbB Wolbachia strain in A. gambiae, and alleviated blood meal-induced mortality in A. stephensi enabling production of Wolbachia-infected offspring. Microbiome analysis using high-throughput sequencing identified that the bacterium Asaia was significantly reduced by antibiotic treatment in both mosquito species. Supplementation of an antibiotic-resistant mutant of Asaia to antibiotic-treated mosquitoes completely inhibited Wolbachia transmission and partly contributed to blood meal-induced mortality. These data suggest that the components of the native mosquito microbiota can impede Wolbachia transmission in Anopheles. Incompatibility between the microbiota and Wolbachia may in part explain why some hosts are uninfected by this endosymbiont in nature.Bacteria in the genus Wolbachia are maternally transmitted Rickettsia-like endosymbionts that infect an estimated 40–69% of arthropod species (1, 2). In many cases, Wolbachia manipulate host reproduction to spread throughout arthropod populations (3). Incongruence between Wolbachia and host phylogenies indicate that horizontal transfer of the symbiont has been commonplace over evolutionary time (4, 5), enabling Wolbachia to invade new species. However, there is a poor understanding of barriers to horizontal transmission and why some species remain uninfected. An understanding of these factors is important from an evolutionary perspective given that Wolbachia influences speciation (6, 7), and from an applied perspective as Wolbachia is being transinfected into vector species for the control of arthropod-borne disease (8–10).The ability to invade the host germ line is an important feature of Wolbachia biology that facilitates horizontal transmission, leading to the pervasive nature of this bacterium across invertebrate taxa. In order for Wolbachia to become established in a naïve host species, it must be acquired horizontally and successfully transmitted vertically (i.e., to offspring) to maintain the infection in the population. Multiple mechanisms of Wolbachia horizontal transmission have been proposed, including cohabitation, hemolymph transfer, predation, and parasitoid infection (11–15). After microinjection into Drosophila, Wolbachia infects the stem cell niches in the germ line (16, 17), and both Wolbachia-derived and host factors appear to influence tropism and bacterial density during oogenesis (17–20). Alternatively, somatic tissue may act as a reservoir for Wolbachia infection of the developing oocyte (20–23). Although pathways of horizontal transmission have been characterized in some species, identification of barriers to vertical transmission of the acquired Wolbachia infection remains elusive.Microbial conflict or incompatibility within arthropods is a potential barrier to transmission of heritable symbionts. Studies in the tick Dermacentor variabilis demonstrate competitive exclusion between maternally inherited bacteria. Transovarial transmission of Rickettsia montanensis (formerly Rickettsia montana) and Rickettsia rhipicephali is inhibited by infection with the reciprocal species (24). Similarly, infection exclusion has been observed in D. variabilis between conspecific strains of Anaplasma marginale where one strain inhibits the infection of the other (25). Competitive inter- and intraspecific microbial interactions have also been observed with Wolbachia (26, 27).Anopheles mosquitoes provide a unique system to examine microbial barriers to Wolbachia transmission. With few exceptions, Anophelines (which transmit the Plasmodium parasites that cause human malaria) are naturally uninfected with Wolbachia (28–31), suggesting the potential presence of innate barriers to infection in this genus. However, in vitro and in vivo studies indicate that Wolbachia are capable of infecting cultured Anopheles cells (32, 33), ex vivo cultured tissues (34), in vivo somatic tissue (35–37), and can stably infect the mosquito germ line (38). We investigated the ability of the native microbial community to influence vertical transmission of Wolbachia in Anopheles mosquitoes. We found that bacteria in the genus Asaia were responsible for inhibiting Wolbachia maternal transmission in this important mosquito genus.     

From the Cover: A holistic picture of Austronesian migrations revealed by phylogeography of Pacific paper mulberry

The peopling of Remote Oceanic islands by Austronesian speakers is a fascinating and yet contentious part of human prehistory. Linguistic, archaeological, and genetic studies have shown the complex nature of the process in which different components that helped to shape Lapita culture in Near Oceania each have their own unique history. Important evidence points to Taiwan as an Austronesian ancestral homeland with a more distant origin in South China, whereas alternative models favor South China to North Vietnam or a Southeast Asian origin. We test these propositions by studying phylogeography of paper mulberry, a common East Asian tree species introduced and clonally propagated since prehistoric times across the Pacific for making barkcloth, a practical and symbolic component of Austronesian cultures. Using the hypervariable chloroplast ndhF-rpl32 sequences of 604 samples collected from East Asia, Southeast Asia, and Oceanic islands (including 19 historical herbarium specimens from Near and Remote Oceania), 48 haplotypes are detected and haplotype cp-17 is predominant in both Near and Remote Oceania. Because cp-17 has an unambiguous Taiwanese origin and cp-17–carrying Oceanic paper mulberries are clonally propagated, our data concur with expectations of Taiwan as the Austronesian homeland, providing circumstantial support for the “out of Taiwan” hypothesis. Our data also provide insights into the dispersal of paper mulberry from South China “into North Taiwan,” the “out of South China–Indochina” expansion to New Guinea, and the geographic origins of post-European introductions of paper mulberry into Oceania.The peopling of Remote Oceania by Austronesian speakers (hereafter Austronesians) concludes the last stage of Neolithic human expansion (1–3). Understanding from where, when, and how ancestral Austronesians bridged the unprecedentedly broad water gaps of the Pacific is a fascinating and yet contentious subject in anthropology (1–8). Linguistic, archaeological, and genetic studies have demonstrated the complex nature of the process, where different components that helped to shape Lapita culture in Near Oceania each have their own unique history (1–3). Important evidence points to Taiwan as an Austronesian ancestral homeland with a more distant origin in South China (S China) (3, 4, 9–12), whereas alternative models suggest S China to North Vietnam (N Vietnam) (7) or a Southeast Asian (SE Asian) origin based mainly on human genetic data (5). The complexity of the subject is further manifested by models theorizing how different spheres of interaction with Near Oceanic indigenous populations during Austronesian migrations have contributed to the origin of Lapita culture (1–3), ranging from the “Express Train” model, assuming fast migrations from S China/Taiwan to Polynesia with limited interaction (4), to the models of “Slow Boat” (5) or “Voyaging Corridor Triple I,” in which “Intrusion” of slower Austronesian migrations plus the “Integration” with indigenous Near Oceanic cultures had resulted in the “Innovation” of the Lapita cultural complex (2, 13).Human migration entails complex skills of organization and cultural adaptations of migrants or colonizing groups (1, 3). Successful colonization to resource-poor islands in Remote Oceania involved conscious transport of a number of plant and animal species critical for both the physical survival of the settlers and their cultural transmission (14). In the process of Austronesian expansion into Oceania, a number of animals (e.g., chicken, pigs, rats, and dogs) and plant species (e.g., bananas, breadfruit, taro, yam, paper mulberry, etc.), either domesticated or managed, were introduced over time from different source regions (3, 8, 15). Although each of these species has been shown to have a different history (8), all these “commensal” species were totally dependent upon humans for dispersal across major water gaps (6, 8, 16). The continued presence of these species as living populations far outside their native ranges represents legacies of the highly skilled seafaring and navigational abilities of the Austronesian voyagers.Given their close association to and dependence on humans for their dispersal, phylogeographic analyses of these commensal species provide unique insights into the complexities of Austronesian expansion and migrations (6, 8, 17). This “commensal approach,” first used to investigate the transport of the Pacific rat Rattus exulans (6), has also been applied to other intentionally transported animals such as pigs, chickens, and the tree snail Partula hyalina, as well as to organisms transported accidentally, such as the moth skink Lipinia noctua and the bacterial pathogen Helicobacter pylori (see refs. 2, 8 for recent reviews).Ancestors of Polynesian settlers transported and introduced a suite of ∼70 useful plant species into the Pacific, but not all of these reached the most isolated islands (15). Most of the commensal plants, however, appear to have geographic origins on the Sahul Plate rather than being introduced from the Sunda Plate or East Asia (16). For example, Polynesian breadfruit (Artocarpus altilis) appears to have arisen over generations of vegetative propagation and selection from Artocarpus camansi that is found wild in New Guinea (18). Kava (Piper methysticum), cultivated for its sedative and anesthetic properties, is distributed entirely to Oceania, from New Guinea to Hawaii (16). On the other hand, ti (Cordyline fruticosa), also a multifunctional plant in Oceania, has no apparent “native” distribution of its own, although its high morphological diversity in New Guinea suggests its origin there (19). Other plants have a different history, such as sweet potato, which is of South American origin and was first introduced into Oceania in pre-Columbian times and secondarily transported across the Pacific by Portuguese and Spanish voyagers via historically documented routes from the Caribbean and Mexico (17).Of all commensal species introduced to Remote Oceania as part of the “transported landscapes” (1), paper mulberry (Broussonetia papyrifera; also called Wauke in Hawaii) is the only species that has a temperate to subtropical East Asian origin (15, 20, 21). As a wind-pollinated, dioecious tree species with globose syncarps of orange–red juicy drupes dispersed by birds and small mammals, paper mulberry is common in China, Taiwan, and Indochina, growing and often thriving in disturbed habitats (15, 20, 21). Because of its long fiber and ease of preparation, paper mulberry contributed to the invention of papermaking in China in A.D. 105 and continues as a prime source for high-quality paper (20, 21). In A.D. 610, this hardy tree species was introduced to Japan for papermaking (21). Subsequently it was also introduced to Europe and the United States (21). Paper mulberry was introduced to the Philippines for reforestation and fiber production in A.D. 1935 (22). In these introduced ranges, paper mulberry often becomes naturalized and invasive (20–22). In Oceania, linguistic evidence suggests strongly an ancient introduction of paper mulberry (15, 20); its propagation and importance across Remote Oceanic islands were well documented in Captain James Cook’s first voyage as the main material for making barkcloth (15, 20).Barkcloth, generally known as tapa (or kapa in Hawaii), is a nonwoven fabric used by prehistoric Austronesians (15, 21). Since the 19th century, daily uses of barkcloth have declined and were replaced by introduced woven textiles; however, tapa remains culturally important for ritual and ceremony in several Pacific islands such as Tonga, Fiji, Samoa, and the SE Asian island of Sulawesi (23). The symbolic status of barkcloth is also seen in recent revivals of traditional tapa making in several Austronesian cultures such as Taiwan (24) and Hawaii (25). To make tapa, the inner bark is peeled off and the bark pieces are expanded by pounding (20, 21, 23). Many pieces of the bark are assembled and felted together via additional poundings to create large textiles (23). The earliest stone beaters, a distinctive tool used for pounding bark fiber, were excavated in S China from a Late Paleolithic site at Guangxi dating back to ∼8,000 y B.P. (26) and from coastal Neolithic sites in the Pearl River Delta dating back to 7,000 y B.P. (27), providing the earliest known archaeological evidence for barkcloth making. Stone beaters dated to slightly later periods have also been excavated in Taiwan (24), Indochina, and SE Asia, suggesting the diffusion of barkcloth culture to these regions (24, 27). These archaeological findings suggest that barkcloth making was invented by Neolithic Austric-speaking peoples in S China long before Han-Chinese influences, which eventually replaced proto-Austronesian language as well as culture (27).In some regions (e.g., Philippines and Solomon Islands), tapa is made of other species of the mulberry family (Moraceae) such as breadfruit and/or wild fig (Ficus spp.); however, paper mulberry remains the primary source of raw material to produce the softest and finest cloth (20, 23). Before its eradication and extinction from many Pacific islands due to the decline of tapa culture, paper mulberry was widely grown across Pacific islands inhabited by Austronesians (15, 20). Both the literature (15, 20) and our own observations (28–30) indicate that extant paper mulberry populations in Oceania are only found in cultivation or as feral populations in abandoned gardens as on Rapa Nui (Easter Island), with naturalization only known from the Solomon Islands (20). For tapa making, its stems are cut and harvested before flowering, and as a majority of Polynesian-introduced crops (16), paper mulberry is propagated clonally by cuttings or root shoots (15, 20), reducing the possibility of fruiting and dispersal via seeds. The clonal nature of the Oceanic paper mulberry has been shown by the lack of genetic variability in nuclear internal transcribed spacer (ITS) DNA sequences (31). With a few exceptions (30), some authors suggest that only male trees of paper mulberry were introduced to Remote Oceania in prehistoric time (15, 20). Furthermore, because paper mulberry has no close relative in Near and Remote Oceania (20), the absence of sexual reproduction precludes the possibility of introgression and warrants paper mulberry as an ideal commensal species to track Austronesian migrations (6, 30).To increase our understanding of the prehistoric Austronesian expansion and migrations, we tracked geographic origins of Oceanic paper mulberry, the only Polynesian commensal plant likely originating in East Asia, using DNA sequence variation of the maternally inherited (32) and hypervariable (SI Text) chloroplast ndhF-rpl32 intergenic spacer (33). We sampled broadly in East Asia (Taiwan, S China, and Japan) and SE Asia (Indochina, the Philippines, and Sulawesi) as well as Oceanic islands where traditional tapa making is still practiced. Historical herbarium collections (A.D. 1899–1964) of Oceania were also sampled to strengthen inferences regarding geographic origins of Oceanic paper mulberry. The employment of ndhF-rpl32 sequences and expanded sampling greatly increased phylogeographic resolution not attainable in a recent study (31) using nuclear ITS sequences (also see SI Text and Fig. S1) and intersimple sequence repeat (ISSR) markers with much smaller sampling.Open in a separate windowFig. S1.ITS haplotype network (n = 17, A–Q) and haplotype distribution and frequency. The haplotype network was reconstructed using TCS (34), with alignment gaps treated as missing data. The sizes of the circles and pie charts are proportional to the frequency of the haplotype (shown in parentheses). Squares denote unique haplotypes (haplotype found only in one individual).     

Physical evidence of predatory behavior in Tyrannosaurus rex

Feeding strategies of the large theropod, Tyrannosaurus rex, either as a predator or a scavenger, have been a topic of debate previously compromised by lack of definitive physical evidence. Tooth drag and bone puncture marks have been documented on suggested prey items, but are often difficult to attribute to a specific theropod. Further, postmortem damage cannot be distinguished from intravital occurrences, unless evidence of healing is present. Here we report definitive evidence of predation by T. rex: a tooth crown embedded in a hadrosaurid caudal centrum, surrounded by healed bone growth. This indicates that the prey escaped and lived for some time after the injury, providing direct evidence of predatory behavior by T. rex. The two traumatically fused hadrosaur vertebrae partially enclosing a T. rex tooth were discovered in the Hell Creek Formation of South Dakota.One of the most daunting tasks of paleontology is inferring the behavior and feeding habits of extinct organisms. Accurate reconstruction of the lifestyle of extinct animals is dependent on the fossil evidence and its interpretation is most confidently predicated on analogy with modern counterparts (1–6). This challenge to understanding the lifestyle of extinct animals is exemplified by the controversy over the feeding behavior of the Late Cretaceous theropod Tyrannosaurus rex (3, 7–17). Although predation and scavenging have often been suggested as distinct feeding behavior alternatives (3, 7–9, 11–17), these terms merit semantic clarification. In this study, predation is considered a subset of feeding behavior, by which any species kills what it eats. Although the term “predator” is used to distinguish such animals from obligate scavengers, it does not imply that the animal did not also scavenge.Ancient diets can be readily reconstructed on the basis of the available evidence, although their derivation (e.g., predation or scavenging behavior) often remains elusive. Speculation as to dinosaur predation has ranged from inferences based on skeletal morphology, ichnofossils such as bite marks, coprolites, stomach contents, and trackways and, by more rarely, direct predator–prey skeletal associations (3, 4, 18–23).Direct evidence of predation in nonavian dinosaurs other than tyrannosaurids has been observed in rare instances, such as the Deinonychus–Tenontosaurus kill site of the Cloverly Formation where the remains of both were found in close association along with shed teeth (9, 24), and the “fighting dinosaurs” from the Gobi Desert, in which a Velociraptor and Protoceratops were found locked in mortal combat (9, 17). The evidence on tyrannosaurids is more limited. Putative stomach contents, such as partially digested juvenile hadrosaur bones, have been reported in association with tyrannosaurid remains (3, 12, 18). This latter instance only represents physical evidence of the last items consumed before the animal’s death, an indicator of diet but not behavior.Mass death assemblages of ornithischians frequently preserve shed theropod teeth (6, 22, 24). Lockley et al. (23) suggest such shed teeth are evidence of scavenging behavior. It is widely argued that T. rex procured food through obligate scavenging rather than hunting (11, 14, 25–27) despite the fact that there is currently no modern analog for such a large bodied obligate scavenger (26). Horner (25) argued that T. rex was too slow to pursue and capture prey items (14) and that large theropods procured food solely through scavenging, rather than hunting (11, 25). Horner also suggested that the enlarged olfactory lobes in T.rex were characteristic of scavengers (25). More recent studies (28, 29) determined the olfactory lobes of modern birds are “poorly developed,” inferring that enlarged olfactory lobes in T. rex are actually a secondary adaptation for predation navigation “to track mobile, dispersed prey” (30). T. rex has a calculated bite force stronger than that of any other terrestrial predator (7), between 35,000 and 57,000 Newtons (30, 31), and possible ambulatory speeds between 20 and 40 kph (7, 15, 16), documenting that it had the capability to pursue and kill prey items.Healed injuries on potential prey animals provide the most unequivocal evidence of survival of a traumatic event (e.g., predation attempt) (3, 32, 33), and several reports attribute such damage to T. rex (4, 17, 19, 20). These include broken and healed proximal caudal vertebral dorsal spines in Edmontosaurus (17) and healed cranial lesions in Triceratops (4, 19). Although the presence of healed injuries demonstrates that an animal lived long enough after the attack to create new bone at the site of the damage (a rare occurrence in the fossil record) (19), the healing usually obliterates any clear signature linking the injury to a specific predator. Bite traces (e.g., raking tooth marks on bone and puncture wounds in the bones of possible prey animals) attributed to T. rex (2, 4, 19) are ambiguous, because the damage inflicted upon an animal during and after a successful hunt mirrors feeding during scavenging. This makes distinction between the two modes of food acquisition virtually impossible with such evidence (3, 34–38).Tooth marks, reported from dinosaur bone-bearing strata worldwide (e.g., 2–4, 8, 19, 20, 39, 40), are further direct evidence of theropod feeding behavior, attributed by some to specific theropod groups (2, 4, 19, 20). Happ (19) and Carpenter (17) identified theropods to family and genus by matching spaces to parallel marks (traces) with intertooth distance. Happ (19) described opposing conical depressions on a left supraorbital Triceratops horn that was missing its distal third (tip), attributing them to a bite by either a T. rex or a crocodilian. Happ (19) stated that the spacing of the parallel marks present on the left squamosal of the same individual matched the intertooth distance of tyrannosaurids. The presence of periosteal reaction documents healing. This contrasts with the report by Farlow and Holtz (3) and again by Hone and Rauhut (20) of the same Hypacrosaurus fibula containing a superficially embedded theropod tooth. Absence of bone reaction precludes confident attribution to predation.Two coalesced hadrosaur (compare with Edmontosaurus annectens) caudal vertebrae were discovered in the Hell Creek Formation of Harding County, South Dakota (40). Archosaur fauna identified in this site include crocodiles, dinosaurs, and birds (41). Physical evidence of dental penetration and extensive infection (osteomylitis) of the fused vertebral centra and healing (bone overgrowth) document an unsuccessful attack by a large predator. A tooth crown was discovered within the wound, permitting identification of the predator as T. rex. This is unambiguous evidence that T. rex was an active predator, fulfilling the criteria that Farlow and Holtz (3) advanced. As T. rex comprises between 1% and 16% of the Upper Cretaceous dinosaurian fauna in Western North America (41–45), its status as a predator or obligate scavenger is nontrivial and could have significant implications for paleoecological reconstructions of that time period. The present contribution provides unique information demonstrating the ecological role for T. rex as that of an active predator. Despite this documentation of predatory behavior by T. rex, we do not make the argument that T. rex was an obligate predator. Like most modern large predators (27, 45) it almost certainly did also scavenge carcasses (9, 16).     

From the Cover: Genomic evidence for inbreeding depression and purging of deleterious genetic variation in Indian tigers

Increasing habitat fragmentation leads to wild populations becoming small, isolated, and threatened by inbreeding depression. However, small populations may be able to purge recessive deleterious alleles as they become expressed in homozygotes, thus reducing inbreeding depression and increasing population viability. We used whole-genome sequences from 57 tigers to estimate individual inbreeding and mutation load in a small–isolated and two large–connected populations in India. As expected, the small–isolated population had substantially higher average genomic inbreeding (F_ROH = 0.57) than the large–connected (F_ROH = 0.35 and F_ROH = 0.46) populations. The small–isolated population had the lowest loss-of-function mutation load, likely due to purging of highly deleterious recessive mutations. The large populations had lower missense mutation loads than the small–isolated population, but were not identical, possibly due to different demographic histories. While the number of the loss-of-function alleles in the small–isolated population was lower, these alleles were at higher frequencies and homozygosity than in the large populations. Together, our data and analyses provide evidence of 1) high mutation load, 2) purging, and 3) the highest predicted inbreeding depression, despite purging, in the small–isolated population. Frequency distributions of damaging and neutral alleles uncover genomic evidence that purifying selection has removed part of the mutation load across Indian tiger populations. These results provide genomic evidence for purifying selection in both small and large populations, but also suggest that the remaining deleterious alleles may have inbreeding-associated fitness costs. We suggest that genetic rescue from sources selected based on genome-wide differentiation could offset any possible impacts of inbreeding depression.

A large proportion of Earth’s biodiversity persists in small and isolated populations in today’s anthropogenically modified world (1). Such populations may suffer from decreased genetic variation and increased inbreeding (2), which together lead to decreased fitness and increased extinction risk (3). Several theoretical (4–7), experimental (8), and empirical studies (9) reveal that species surviving in small and isolated populations are at the greatest risk of extinction.While species exist in nature along a continuum from small to large populations with different levels of isolation, populations of endangered species often tend to be small and isolated. African wild dog, Ethiopian wolf, and great Indian bustard are examples of species where all populations are small and isolated (10–12). The “small population paradigm” of conservation biology suggests that such smaller and more isolated populations are at a higher risk of extinction due to inbreeding depression and demographic stochasticity (13–15).Inbred individuals express deleterious, partially recessive alleles that are inherited identically by descent (IBD) from related parents, leading to inbreeding depression (16). Such inbreeding depression can reduce the average fitness of a population, eventually leading to reduced population size and possibly extinction (17). A commonly adopted strategy to conserve inbred populations is genetic rescue (18), which aims to increase average fitness by decreasing the frequency of deleterious mutations and increase heterozygosity at loci harboring deleterious alleles, via translocations of individuals from genetically differentiated populations. A meta-analysis of empirical data from wild populations showed broadly consistent positive effects of genetic rescue on fitness (15, 19).Population genetic theory (20–22) predicts that purifying selection can reduce inbreeding depression by purging deleterious alleles from inbred populations in the absence of immigration. Whether isolated populations are likely to purge a substantial fraction of the mutation load has been of longstanding interest in evolutionary biology and conservation. Early empirical data from pedigreed captive populations suggested that purging either was absent or resulted only in slight decreases in inbreeding depression (23, 24). However, several experimental studies based on model organisms reveal substantial purging and significant reduction in inbreeding depression in small populations (25–27). Recent molecular and population genetic studies have found genomic evidence for purging in wild populations (28–30). Despite broad empirical support for the efficacy of genetic rescue (15, 19), genomic evidence for purging and the long-term persistence of some small–isolated populations have been cited to question the small population paradigm and to argue that standard genetic rescue practices are likely to be counterproductive (29, 31–33). Whether purging removes enough deleterious alleles to improve the viability of small, isolated populations (contradicting the small population paradigm) remains an open question. We address this question by contrasting genomic inbreeding and mutation load in small–isolated versus large–connected populations of wild tigers.We use Bengal tigers (Panthera tigris tigris) from India as a model to investigate levels of inbreeding and relative mutation loads in small–isolated and large–connected populations and examine the potential for genetic rescue. Tigers are large, endangered carnivores, but Bengal tigers have high genetic variation compared to other subspecies, with some subpopulations showing high inbreeding indicating isolation (34). All Bengal tiger populations have been through historic bottlenecks, but inbreeding and genetic variation vary among populations. Also, some populations are relatively large and connected while others are small and isolated from both genetic (35) and demographic perspectives (36, 37), making them an ideal system to investigate inbreeding, mutation load, and possible genetic rescue strategies.We use genomic data to measure the impact of historically declining population sizes and connectivity on inbreeding and mutation load in three wild Bengal tiger (P. tigris tigris) populations that are small–isolated (SI) and large–connected (LC). The small–isolated population is from northwestern India and the large–connected populations are from southern India (s-LC) and central India (c-LC). We expected large–connected populations to be the least inbred and to have the lowest mutation load. Alternatively, purging could result in lower mutation load in this small–isolated population. We also explore strategies for genetic rescue that might effectively decrease inbreeding depression. For tigers, we specifically suggest strategies for identifying populations that may benefit from genetic rescue and how such strategies may be effectively implemented.     

Density-dependent mating behaviors reduce male mating harassment in locusts

Male mating harassment may occur when females and males do not have the same mating objectives. Communal animals need to manage the costs of male mating harassment. Here, we demonstrate how desert locusts in dense populations reduce such conflicts through behaviors. In transient populations (of solitarious morphology but gregarious behavior), we found that nongravid females occupied separate sites far from males and were not mating, whereas males aggregated on open ground (leks), waiting for gravid females to enter the lekking sites. Once a male mounted a gravid female, no other males attacked the pair; mating pairs were thereby protected during the vulnerable time of oviposition. In comparison, solitarious locusts displayed a balanced sex ratio in low-density populations, and females mated irrespective of their ovarian state. Our results indicate that the mating behaviors of desert locusts are density dependent and that sex-biased behavioral group separation may minimize the costs of male mating harassment and competition.

The maintenance of animal societies depends on conflict management (1–3). Sexual interaction among group members frequently causes competition and male mating harassment (4). Males typically attempt to maximize fitness by fertilizing as many females as possible (5). Consequently, females are frequently harassed by males, with severe male–male competition (6). Communal animals need to manage the costs of male mating harassment as well as male–male competition.The operational sex ratio (OSR) is usually biased toward males because males tend to recover their reproductive state faster than females (7). Male-biased OSRs cause male–male competition, leading to intense sexual coercion of females by males (i.e., male mating harassment). This phenomenon reduces female fitness by increasing the risk of predation (8) and injury (9). In such cases, a sexually antagonistic coevolution, or “arms race,” of male persistence versus female resistance may occur. This is termed “sexual conflict” (10, 11). Sexual conflicts occur when females and males do not have the same mating objectives (12, 13). However, this phenomenon generates substantial costs, resulting in the loss of energy, time, and mating chance (14). It may also increase predation risk (12), which may negatively influence population dynamics (15). Exaggerated arms races may be counterproductive for communal animals. OSR theory proposes the following simple solution for how animals can resolve these opposing forces: females should live separately from males to prevent male mating harassment (16). According to this theory, nongravid females should occupy sites without males (“time out”), whereas females that are gravid and ready for oviposition should enter male-biased groups (the mating pool) for mating and oviposition (“time in”). In some species, males may form leks (i.e., aggregations), where they engage in competitive displays to entice visiting females that seek partners for copulation (2, 4, 17). The evolutionary mechanism explaining these paradoxical lekking behaviors remains controversial (2, 17). Several competing and nonexclusive models have been proposed by considering various factors including predation risk, habitat fragmentation, resource distribution, and male mating harassment (18). In some cases, males are directly or indirectly evaluated based on their quality by females investing in mate choice (17, 18). However, information about whether this system has evolved in communal animals is limited, especially for species with density-dependent behaviors. Here, we explored how the desert locust, Schistocerca gregaria, resolves male mating harassment and male–male competition in the field.Desert locusts change from solitary to group life plastically; that is, they change from an initial “solitarious phase,” during which they do not aggregate, to a “gregarious phase,” during which they form swarms that migrate long distances (19, 20). This extreme phenotypic plasticity (termed phase polyphenism) depends on local population density (21, 22), and the associated behavioral and physiological traits may change quickly during the life cycle (23, 24). A behavioral difference is measurable within 1 h and almost completed within 4 h (23). In contrast, other traits such as adult morphometric ratios do not change after adult eclosion. Thus, solitarious-phase looking adult locusts may behave as typical gregarious locusts [referred to as transient (25) or gregarious-behaving locusts; for an explanation of terms, see Study Animal and Terms Used to Describe Populations]. For gregarious-behaving locusts, little is known about how nongravid females prevent mating harassment by males. Although the desert locust is a major pest in over 60 countries in Africa and Asia (26), its mating strategies and management of male mating harassment at the group level remain unclear (19, 27).During field observations over 9 y in the Sahara Desert of Mauritania, we noticed biased sex ratios of sexually mature groups of desert locusts during the transition from the solitarious to the gregarious phase. In female-biased groups, most females were single (i.e., not guarded by a male), whereas most females were mated in male-biased groups. Researchers previously reported that sexually mature swarms tend to split after oviposition but frequently rejoin when they resume migratory flight between successive oviposition cycles (27). These fragmentary observations were consistent with the “group separation” mating system predicted by the OSR theory (16). Based on these observations and theory, we hypothesized that sexually mature, gregarious-behaving desert locust females and males occupied separate areas (depending on the state of ovarian development) to prevent male mating harassment and to offset costs of male–male competition in mating with gravid females.We tested this hypothesis by surveying gregarious-behaving populations of desert locusts in the field and by conducting parallel laboratory experiments. Because mating systems vary depending on population density (28), we also examined the mating system of solitarious-phase locusts in low-density populations. Here, we show how desert locusts use a previously unrecognized density-dependent “group separation” mating system to manage male mating harassment and competition without losing sociability.     

From the Cover: Enlisting wild grass genes to combat nitrification in wheat farming: A nature-based solution

Active nitrifiers and rapid nitrification are major contributing factors to nitrogen losses in global wheat production. Suppressing nitrifier activity is an effective strategy to limit N losses from agriculture. Production and release of nitrification inhibitors from plant roots is termed “biological nitrification inhibition” (BNI). Here, we report the discovery of a chromosome region that controls BNI production in “wheat grass” Leymus racemosus (Lam.) Tzvelev, located on the short arm of the “Lr#3Ns^b” (Lr#n), which can be transferred to wheat as T3BL.3Ns^bS (denoted Lr#n-SA), where 3BS arm of chromosome 3B of wheat was replaced by 3Ns^bS of L. racemosus. We successfully introduced T3BL.3Ns^bS into the wheat cultivar “Chinese Spring” (CS-Lr#n-SA, referred to as “BNI-CS”), which resulted in the doubling of its BNI capacity. T3BL.3Ns^bS from BNI-CS was then transferred to several elite high-yielding hexaploid wheat cultivars, leading to near doubling of BNI production in “BNI-MUNAL” and “BNI-ROELFS.” Laboratory incubation studies with root-zone soil from field-grown BNI-MUNAL confirmed BNI trait expression, evident from suppression of soil nitrifier activity, reduced nitrification potential, and N₂O emissions. Changes in N metabolism included reductions in both leaf nitrate, nitrate reductase activity, and enhanced glutamine synthetase activity, indicating a shift toward ammonium nutrition. Nitrogen uptake from soil organic matter mineralization improved under low N conditions. Biomass production, grain yields, and N uptake were significantly higher in BNI-MUNAL across N treatments. Grain protein levels and breadmaking attributes were not negatively impacted. Wide use of BNI functions in wheat breeding may combat nitrification in high N input–intensive farming but also can improve adaptation to low N input marginal areas.

Nitrification and denitrification are critical soil biological processes, which, left unchecked, can accelerate generation of harmful reactive nitrogen (N) forms (NO₃ ⁻, N₂O, and NOx) that trigger a “nitrogen cascade,” damaging ecosystems, water systems, and soil fertility (1 –8). Excessive nitrifier activity and a rapid generation of soil nitrates plague modern cereal production systems. This has led to shifting crop N nutrition toward an “all nitrate form,” which is largely responsible for N losses and a decline in agronomic nitrogen-use efficiency (NUE) (6, 7, 9 –11).Wheat, one of the three founding crops for food security (12), consumes nearly a fifth of factory-produced N fertilizers, and it has an average NUE of 33%, which has remained unchanged for the last two decades (13 –15). Regulating soil nitrifier activity to slow the rate of soil nitrate formation should provide more balanced N forms (NH₄ ⁺ and NO₃ ⁻) for plant uptake (rather than nearly “all NO₃ ⁻” at present), reduce N losses, and facilitate the assimilation of dual N forms. This optimizes the utilization of biochemical machinery for N assimilation, improving stability and possibly enhancing yield potential (16). In addition, the assimilation of NH₄ ⁺ is energetically more efficient (requiring 40% less metabolic energy) than NO₃ ⁻ assimilation (16). Often, a stimulatory growth response is observed in wheat, when 15 to 30% of NO₃ ⁻ is replaced with NH₄ ⁺ in nutrient solutions (17, 18).Synthetic nitrification inhibitors (SNIs) have been shown to suppress N₂O emissions, reduce N losses, and improve agronomic NUE in several cereal crops including wheat (6, 19 –21). However, the lack of cost effectiveness, inconsistency in field performance, inability to function in tropical environments, and the concerns related to the entering of SNIs into food chains have limited their adoption in production agriculture (6, 7, 19, 20).Biological nitrification inhibition (BNI) is a plant function whereby nitrification inhibitors (BNIs) are produced from root systems to suppress soil nitrifier activity (22 –26). Earlier, we reported that the BNI capacity in the root systems of cultivated wheat lack adequate strength to effectively suppress soil nitrifier activity in the rhizosphere (24, 25). Leymus racemosus (hereafter referred to as “wild grass”), a perennial Triticeae evolutionarily related to wheat, produces extensive root systems ( SI Appendix, Fig. S1) and was discovered to have a high BNI capacity several times higher than cultivated wheat. It was also effective in suppressing soil nitrifier activity and in reducing soi -nitrate formation ( SI Appendix, Fig. S2) (25). Subsequently, the chromosome Lr#n = 3Ns^b was found to be controlling a major part of BNI capacity in wild grass, and it is the focus of our current research (25, 27, 28). Earlier, we reported that Lr#I and Lr#J had a minor impact on BNI capacity, but they are not the focus of this research (25).We transferred the Lr#n chromosome (Lr#n-SA = T3BL.3Ns^bS) controlling BNI capacity (hereafter referred to as BNI trait) into the cultivated wheat, Chinese Spring (CS). The results of the transfer of this BNI trait into several elite wheat types with a grain-yield (GY) potential >10 t ha⁻¹, resulting in substantial improvements of BNI capacity in root systems, are reported in this paper.     

Link recommendation algorithms and dynamics of polarization in online social networks

The level of antagonism between political groups has risen in the past years. Supporters of a given party increasingly dislike members of the opposing group and avoid intergroup interactions, leading to homophilic social networks. While new connections offline are driven largely by human decisions, new connections on online social platforms are intermediated by link recommendation algorithms, e.g., “People you may know” or “Whom to follow” suggestions. The long-term impacts of link recommendation in polarization are unclear, particularly as exposure to opposing viewpoints has a dual effect: Connections with out-group members can lead to opinion convergence and prevent group polarization or further separate opinions. Here, we provide a complex adaptive–systems perspective on the effects of link recommendation algorithms. While several models justify polarization through rewiring based on opinion similarity, here we explain it through rewiring grounded in structural similarity—defined as similarity based on network properties. We observe that preferentially establishing links with structurally similar nodes (i.e., sharing many neighbors) results in network topologies that are amenable to opinion polarization. Hence, polarization occurs not because of a desire to shield oneself from disagreeable attitudes but, instead, due to the creation of inadvertent echo chambers. When networks are composed of nodes that react differently to out-group contacts, either converging or polarizing, we find that connecting structurally dissimilar nodes moderates opinions. Overall, our study sheds light on the impacts of social-network algorithms and unveils avenues to steer dynamics of radicalization and polarization in online social networks.

Online social networks are increasingly used to access political information (1), engage with political elites, and discuss politics (2). These new communication platforms can benefit democratic processes in several ways: They reduce barriers to information and, subsequently, increase citizen engagement, allow individuals to voice their concerns, help debunk false information, and improve accountability and transparency in political decision-making (3). In principle, individuals can use social media to access ideologically diverse viewpoints and make better-informed decisions (4, 5).At the same time, internet and online social networks reveal a dark side. There are mounting concerns over possible linkages between social media and affective polarization (6, 7). Other than healthy political deliberation, social networks can foster so-called “echo chambers” (8, 9) and “information cocoons” (3, 10) where individuals are only exposed to like-minded peers and homogeneous sources of information, which polarizes attitudes (for counterevidence, see ref. 5). As a result, social media can trigger political sectarianism (6, 7, 11–13) and fuel misinformation (14, 15). Averting the risks of online social networks for political institutions, and potentiating their advantages, requires multidisciplinary approaches and novel methods to understand long-term dynamics on social platforms.That is not an easy task. As pointed out by Woolley and Howard, “to understand contemporary political communication we must now investigate the politics of algorithms and automation” (16). While traditional media outlets are curated by humans, online social media resorts to computer algorithms to personalize contents through automatic filtering. To understand information dynamics in online social networks, one needs to take into account the interrelated subtleties of human decision making [e.g., only share specific contents (17), actively engage with other users, follow or befriend particular individuals, interact offline] and the outcomes of automated decisions (e.g., news sorting and recommendation systems) (18, 19). In this regard, much attention has been placed on the role of news filters and sorting (1, 18, 19). Shmargad and Klar (20) provide evidence that algorithms sorting news impact the way users engage with and evaluate political news, likely exacerbating political polarization. Likewise, Levy (21) notes that social media algorithms can substantially affect users’ news consumption habits.While past studies have examined how algorithms may affect which information appears on a person’s newsfeed, and subsequent polarization, social matching (22) or link recommendation (23) algorithms [also called user, contact, or people recommender systems (24, 25)] constitute another class of algorithms that can affect the way users engage in (and with) online social networks (examples of such systems in SI Appendix, Fig. S13). These algorithms are implemented to recommend new online connections—“friends” or “followees”—to social network users, based on supposed offline familiarity, likelihood of establishing a future relation, similar interests, or the potential to serve as a source of useful information. Current data provide evidence that link recommendation algorithms impact network topologies and increase network clustering: Daly et al. (26) show that an algorithm recommending friends-of-friends, in an IBM internal social network platform, increases clustering and network modularity. Su et al. (27) analyzed the Twitter graph before and after this platform implemented link recommendation algorithms and show that the “Who To Follow” feature led to a sudden increase in edge growth and the network clustering coefficient. Similarly, Zignani et al. (28) show that, on a small sample of the Facebook graph, the introduction of the “People You May Know” (PYMK) feature led to a sudden increase in the number of links and triangles [i.e., motifs comprising three nodes (A, B, C) where the links AB, AC, and BC exist] in the network. The fact that PYMK is responsible for a significant fraction of link creations is alluded to in other works (29). Furthermore, recent work shows, through experiments with real social media users (30) and simulations (31), that link recommendation algorithms can effectively be used as an intervention mechanism to increase networks’ structural diversity (30, 31) and minimize disagreements (32). It is thereby relevant to understand, 1) How do algorithmic link recommendations interplay with opinion formation? and 2) What are the long-term impacts of such algorithms on opinion polarization?Here, we tackle the previous questions from a complex adaptive–systems perspective (33), designing and analyzing a simple model where individuals interact in a dynamic social network. While several models explain the emergence of polarization through link formation based on opinion similarity (34–41) and information exchange (42), here we focus instead on rewiring based on “structural similarity,” which is defined as similarity based on common features that exclusively depend on the network structure (43). This contrasts with the broader concept of homophily, which typically refers to similarity based on common characteristics besides network properties (e.g., opinions, taste, age, background). Compared with rewiring based on homophily—which can also contribute to network fragmentation—rewiring based on structural similarity can be less restrictive in contexts where information about opinions and beliefs is not readily available to individuals before the connection is established. Furthermore, rewiring based on structural similarity is a backbone of link recommendation algorithms [e.g., “People you may know” or “Whom to follow” (25) suggestions], which rely on link prediction methods to suggest connections to users (43, 44). Importantly, our model combines three key ingredients: 1) Links are formed according to structural similarity, based on common neighbors, which is one of the simplest link prediction methods (43); this way, we do not assume a priori that individuals with similar opinions are likely to become connected [as recent works underline, sorting can be incidental to politics (45, 46)]. 2) Then, to examine opinion updating, we adapt a recent model that covers the interplay of social reinforcement and issue controversy to promote radicalization on social networks (39). 3) Last, we explicitly consider that nodes can react differently to out-group links, either converging in their opinions (10, 47) or polarizing further (48–50).We find that establishing links based on structural similarity alone [a process that is likely to be reinforced by link recommendation algorithms—see SI Appendix, Fig. S10 and previous work pointing that such algorithms affect a social network topology and increase their clustering coefficient (26–28)] contributes to opinion polarization. While our model sheds light on the effect of link recommendation algorithms on opinion formation and polarization dynamics, we also offer a justification for polarization to emerge through structural similarity-based rewiring, in the absence of explicit opinion-similarity rewiring (34, 36, 39, 51), confidence-bounds (37, 38, 40), or rewiring based on concordant messages (42).^* Second, we find that the effects of structural similarity-based rewiring are exacerbated if even moderate opinions have high social influence. Finally, we combine nodes that react differently to out-group contacts: “converging” nodes, which converge if exposed to different opinions (10, 21, 52), and “polarizing” nodes, which diverge when exposed to different viewpoints (48–50). We observe that the coexistence of both types of nodes can contribute to moderate opinions. Polarizing nodes develop radical opinions, and converging nodes, influenced by opposing viewpoints, yield more temperate ones. However, again, link recommendation algorithms impact this process: Given the existence of communities isolated to a greater degree through link recommendation, converging nodes may find it harder to access diverse viewpoints, which, in general, contributes to increasing the adoption of extreme opinions.     

Obstacles to integrated pest management adoption in developing countries

Despite its theoretical prominence and sound principles, integrated pest management (IPM) continues to suffer from anemic adoption rates in developing countries. To shed light on the reasons, we surveyed the opinions of a large and diverse pool of IPM professionals and practitioners from 96 countries by using structured concept mapping. The first phase of this method elicited 413 open-ended responses on perceived obstacles to IPM. Analysis of responses revealed 51 unique statements on obstacles, the most frequent of which was “insufficient training and technical support to farmers.” Cluster analyses, based on participant opinions, grouped these unique statements into six themes: research weaknesses, outreach weaknesses, IPM weaknesses, farmer weaknesses, pesticide industry interference, and weak adoption incentives. Subsequently, 163 participants rated the obstacles expressed in the 51 unique statements according to importance and remediation difficulty. Respondents from developing countries and high-income countries rated the obstacles differently. As a group, developing-country respondents rated “IPM requires collective action within a farming community” as their top obstacle to IPM adoption. Respondents from high-income countries prioritized instead the “shortage of well-qualified IPM experts and extensionists.” Differential prioritization was also evident among developing-country regions, and when obstacle statements were grouped into themes. Results highlighted the need to improve the participation of stakeholders from developing countries in the IPM adoption debate, and also to situate the debate within specific regional contexts.Feeding the 9,000 million people expected to inhabit Earth by 2050 will present a constant and significant challenge in terms of agricultural pest management (1–3). Despite a 15- to 20-fold increase in pesticide use since the 1960s, global crop losses to pests—arthropods, diseases, and weeds—have remained unsustainably high, even increasing in some cases (4). These losses tend to be highest in developing countries, averaging 40–50%, compared with 25–30% in high-income countries (5). Alarmingly, crop pest problems are projected to increase because of agricultural intensification (4, 6), trade globalization (7), and, potentially, climate change (8).Since the 1960s, integrated pest management (IPM) has become the dominant crop protection paradigm, being endorsed globally by scientists, policymakers, and international development agencies (2, 9–15). The definitions of IPM are numerous, but all involve the coordinated integration of multiple complementary methods to suppress pests in a safe, cost-effective, and environmentally friendly manner (9, 11). These definitions also recognize IPM as a dynamic process in terms of design, implementation, and evaluation (11). In practice, however, there is a continuum of interpretations of IPM (e.g., refs. 14, 16, 17), but bounded by those that emphasize pesticide management (i.e., “tactical IPM”) and those that emphasize agroecosystem management (i.e., “strategic IPM,” also known as “ecologically based pest management”) (16, 18, 19). Despite apparently solid conceptual grounding and substantial promotion by the aforementioned groups, IPM has a discouragingly poor adoption record, particularly in developing-country settings (9, 10, 15–23), raising questions over its applicability as it is presently conceived (15, 16, 22, 24).The possible reasons behind the developing countries’ poor adoption of IPM have been the subject of considerable discussion since the 1980s (9, 15, 16, 22, 25–31), but this debate has been notable for the limited direct involvement from developing-country stakeholders. Most of the literature exploring poor adoption of IPM in the developing world has originated in the developed world (e.g., refs. 15, 16, 22). An international workshop, entitled “IPM in Developing Countries,” was held at the Pontificia Universidad Católica del Ecuador (PUCE) from October 31 to November 3, 2011. Poor IPM adoption spontaneously became a central discussion point, creating an opportunity to address the apparent participation bias in the IPM adoption debate.It was therefore decided to explore the topic further by eliciting and mapping the opinions of a large and diverse pool of IPM professionals and practitioners from around the world, including many based in developing countries. The objective was to generate and prioritize a broad list of hypotheses to explain poor IPM adoption in developing-country agriculture. We also wanted to explore differences as influenced by respondents’ characteristics, particularly their region of practice. To achieve these objectives, we used structured concept mapping (32), an empirical survey method often used to quantify and give thematic structure to open-ended opinions (33).We know of only one other similar study that characterizes obstacles to IPM. It was based on the structured responses of 153 experts, all from high-income countries (30). Our survey was designed to progress from unstructured to structured responses, and to reach a much larger and diverse pool of participants, particularly those from the “Global South.” Considering that the vast majority of farmers live in developing countries (34), it would seem imperative that the voices from this region be heard.     

Metapopulations with habitat modification

Across the tree of life, organisms modify their local environment, rendering it more or less hospitable for other species. Despite the ubiquity of these processes, simple models that can be used to develop intuitions about the consequences of widespread habitat modification are lacking. Here, we extend the classic Levins metapopulation model to a setting where each of n species can colonize patches connected by dispersal, and when patches are vacated via local extinction, they retain a “memory” of the previous occupant—modeling habitat modification. While this model can exhibit a wide range of dynamics, we draw several overarching conclusions about the effects of modification and memory. In particular, we find that any number of species may potentially coexist, provided that each is at a disadvantage when colonizing patches vacated by a conspecific. This notion is made precise through a quantitative stability condition, which provides a way to unify and formalize existing conceptual models. We also show that when patch memory facilitates coexistence, it generically induces a positive relationship between diversity and robustness (tolerance of disturbance). Our simple model provides a portable, tractable framework for studying systems where species modify and react to a shared landscape.

Many interactions between species are realized indirectly, through effects on a shared environment. For example, consumers compete indirectly by altering resource availability (1, 2). However, the ways that species affect and are affected by their environment extend far beyond the consumption of resources. Across the tree of life and over a tremendous range of spatial scales, organisms make complex and sometimes substantial changes to the physical and chemical properties of their local environment (3–6). Many species also impact local biotic factors; for example, plant–soil feedbacks are often driven by changes in soil microbiome composition (4, 7–9).Numerous studies have recognized and discussed the ways that such changes can mediate interactions between species, as well as the obstacles to modeling these complex, indirect interactions (5, 7, 10–12). In some instances, the effects of environmental modification by one species on another can be accounted for implicitly in models of direct interactions (2, 13, 14) or within the well-established framework of resource competition (12, 15). But in many other cases, new modeling approaches are necessary.Because the range of ecosystems where interactions are driven by environmental modification is wide and varied, many parallel strands of theory have developed for them. Examples include “traditional” ecosystem engineers (16–20), plant–soil feedbacks (4, 7, 21), and chemically mediated interactions between microbes (5, 12). Similar dynamics underlie Janzen–Connell effects, where individuals (e.g., tropical trees) modify their local environment by supporting high densities of natural enemies (8, 22–24), and immune-mediated pathogen competition, where pathogen strains modify their hosts by inducing specific immunity (25–28). These last two examples highlight that environmental modification might be “passive,” in the sense that it is generated by the environment itself.While each of these systems has attracted careful study, it is difficult to elucidate general principles for the dynamics of environmentally mediated interactions without a simple, shared theoretical framework. Are there generic conditions for the coexistence of many species in these systems? What are typical relationships between diversity and ecosystem productivity or robustness? We especially lack theoretical expectations for high-diversity communities, as most existing models focus on the dynamics of one or two species (4, 7, 16, 17).To begin answering these questions, we introduce and analyze a flexible model for species interactions mediated by environmental modification. Two essential features of these interactions—which underlie the difficulty integrating them into standard ecological theory—are that environmental modifications are localized in space and persistent in time (10). To capture these aspects, we adopt the metapopulation framework, introduced by Levins (29), which provides a minimal model for population dynamics with distinct local and global scales. Metapopulation models underpin a productive and diverse body of theory in ecology (30, 31), including various extensions to study multispecies communities (32, 33). Here, we adopt the simplest such extension, by assuming zero-sum dynamics and an essentially horizontal community (34, 35). Our modeling framework accommodates lasting environmental modification by introducing a versatile notion of “patch memory,” in which the state of local sites depends on past occupants.In line with evidence from a range of systems, we find that patch memory can support the robust coexistence of any number of species, even in an initially homogeneous landscape. We derive quantitative conditions for species’ coexistence and show how they connect to existing conceptual models. Importantly, these conditions apply even as several model assumptions are relaxed. We also investigate an emergent relationship between species diversity and robustness, demonstrating that our modeling framework can provide insight for a variety of systems characterized by localized environmental feedbacks.     

Cyclodextrins increase membrane tension and are universal activators of mechanosensitive channels

The bacterial mechanosensitive channel of small conductance (MscS) has been extensively studied to understand how mechanical forces are converted into the conformational changes that underlie mechanosensitive (MS) channel gating. We showed that lipid removal by β-cyclodextrin can mimic membrane tension. Here, we show that all cyclodextrins (CDs) can activate reconstituted Escherichia coli MscS, that MscS activation by CDs depends on CD-mediated lipid removal, and that the CD amount required to gate MscS scales with the channel’s sensitivity to membrane tension. Importantly, cholesterol-loaded CDs do not activate MscS. CD-mediated lipid removal ultimately causes MscS desensitization, which we show is affected by the lipid environment. While many MS channels respond to membrane forces, generalized by the “force-from-lipids” principle, their different molecular architectures suggest that they use unique ways to convert mechanical forces into conformational changes. To test whether CDs can also be used to activate other MS channels, we chose to investigate the mechanosensitive channel of large conductance (MscL) and demonstrate that CDs can also activate this structurally unrelated channel. Since CDs can open the least tension-sensitive MS channel, MscL, they should be able to open any MS channel that responds to membrane tension. Thus, CDs emerge as a universal tool for the structural and functional characterization of unrelated MS channels.

Bacterial mechanosensitive (MS) channels have been extensively used as models of ion channel–mediated mechanotransduction (1, 2). They have continually provided novel insights into the biophysical principles that govern ion-channel mechanosensitivity (3–6). While the structurally unrelated MS channels MscL (mechanosensitive channel of large conductance) (4) and MscS (mechanosensitive channel of small conductance) (7) both respond to changes in membrane tension (8–10), at the molecular level, they seem to employ different strategies to convert membrane forces into the conformational changes that underlie channel gating.Escherichia coli MscS is the archetypal member of a large structurally diverse family of ion channels that are expressed in bacteria (11, 12), archaea (13), some fungi (14), plants (15, 16), and eukaryotic parasites (17). This channel gates as a result of membrane tension (10) in accordance with the “force-from-lipids” gating mechanism (6). In response to increases in membrane tension, MscS exhibits complex adaptive gating kinetics (18–20). These kinetic responses may represent two separable processes, adaptation and inactivation (21, 22). In particular, point mutations within transmembrane domain 3 can instigate phenotypes in which adaptation and inactivation are affected differently (18, 23, 24). These complex kinetics are important for the role of this channel as an osmotic safety valve (25). However, since it is currently unknown whether these electrophysiologically separable processes correlate to structurally distinct states, we will refer to them collectively as “desensitization.” In addition, while some data suggest that MscS desensitization is sensitive to the lipid environment (26), this notion still awaits definitive proof.MscL was the first MS channel to be cloned and functionally characterized in a lipid-only environment (8). Members of the MscL family, unlike those of the MscS family, are almost exclusively expressed in archaea and bacteria. After X-ray crystallography revealed the structure of MscL (27), subsequent studies implicated membrane thinning in response to membrane tension as a major driver of MscL gating (3, 28).To fully understand the structural basis of the gating transitions in MscL and MscS, one must first find a way to apply a gating stimulus to the channels in a lipidic environment that is compatible with structural studies. This is, of course, less challenging when considering ligand-gated channels (29–31), for which the stimulus is a defined molecule that can readily be applied to visualize the resulting changes in protein conformation. For MS channels, until recently, only spectroscopic approaches, such as electron paramagnetic resonance spectroscopy (32, 33) and Förster resonance energy transfer spectroscopy (34, 35), were available to provide structural insights into their gating in response to changes in forces in their lipid environment. Other approaches had been confined to the use of activators (36) or mutations (37, 38). We recently demonstrated that lipid removal by β-cyclodextrin can mimic membrane tension in membrane-scaffold protein-based lipid nanodiscs, providing novel insights into the structural rearrangements that underlie MscS channel gating in response to membrane tension (39). The idea was that, as long as the surface area would not change and the lipids would not be replaced, β-cyclodextrin–mediated lipid removal from a membrane would result in the remaining lipids having to cover a larger surface area. This increase in “area-per-lipid” would result in a corresponding increase in membrane tension (40, 41) that would be experienced by integral membrane proteins incorporated in that membrane.Cyclodextrins (CDs) are a family of cyclic glucose oligomers with a cone-like three-dimensional architecture characterized by a polar external surface and a hydrophobic cavity (42, 43). α-, β-, and γ-CD contain six to eight glucose units, respectively. As the number of units increases, so does the diameter of the hydrophobic cavity (5 to 8 Å) (43). These compounds are of broad utility, as the hydrophobic cavity can chelate a plethora of small lipophilic molecules (44, 45). CDs can also form complexes with fatty acids and phospholipids (46). CDs have thus been widely used to remove lipids from native cell membranes (47, 48) and from model membranes (49, 50). This removal of lipids has already been directly linked to increases in membrane tension even in intact cellular environments (51). CDs also exhibit differential lipid selectivity. For example, α-CD has the selectivity profile of phosphatidylserine > phosphatidylethanolamine >> phosphatidylcholine (52). The methylated version of β-CD (mβ-CD) shows selectivity toward cholesterol at low concentrations and has been widely used to selectively remove or add cholesterol to cell membranes (48, 53–55). In addition to the headgroup, CDs also preferentially chelate unsaturated lipids and those containing shorter acyl chains (56, 57).Here, we show that all members of the CD family (α, β, and γ) can activate E. coli MscS in liposomal membranes. Even the methylated version of β-CD, which is widely used for its cholesterol selectivity, can activate E. coli MscS. Congruent with lipid removal increasing tension, the CD amount required for the activation of an MS channel depends on its tension sensitivity. Importantly, as a control, cholesterol-loaded CDs do not activate MscS. Our studies also clearly establish that MscS desensitization is modified by the lipid environment. Moreover, we show that CD-mediated lipid removal causes a concentration- and time-dependent increase in the tension in excised membrane patches and that the resulting tension can become sufficiently high to activate the structurally unrelated MS channel MscL that gates at membrane tensions immediately below the lytic limit of membranes. Two-dimensional (2D) class averages of nanodisc-embedded MscL obtained by cryo–electron microscopy (cryo-EM) indicate that β-CD treatment results in membrane thinning and channel expansion. The fact that CD activates MscL, which opens immediately below the lytic tension of the membrane, suggests that all other MS channels (which are all more sensitive to membrane tension) should also open in response to CDs. These data suggest that CDs will be of broad utility for the structural and functional characterization of structurally diverse MS channels, including Piezo channels (58, 59), two-pore domain K⁺ channels (40, 60), and OSCA channels (61, 62), all of which are known to respond to membrane forces.     

An architecture for encoding sentence meaning in left mid-superior temporal cortex

Human brains flexibly combine the meanings of words to compose structured thoughts. For example, by combining the meanings of “bite,” “dog,” and “man,” we can think about a dog biting a man, or a man biting a dog. Here, in two functional magnetic resonance imaging (fMRI) experiments using multivoxel pattern analysis (MVPA), we identify a region of left mid-superior temporal cortex (lmSTC) that flexibly encodes “who did what to whom” in visually presented sentences. We find that lmSTC represents the current values of abstract semantic variables (“Who did it?” and “To whom was it done?”) in distinct subregions. Experiment 1 first identifies a broad region of lmSTC whose activity patterns (i) facilitate decoding of structure-dependent sentence meaning (“Who did what to whom?”) and (ii) predict affect-related amygdala responses that depend on this information (e.g., “the baby kicked the grandfather” vs. “the grandfather kicked the baby”). Experiment 2 then identifies distinct, but neighboring, subregions of lmSTC whose activity patterns carry information about the identity of the current “agent” (“Who did it?”) and the current “patient” (“To whom was it done?”). These neighboring subregions lie along the upper bank of the superior temporal sulcus and the lateral bank of the superior temporal gyrus, respectively. At a high level, these regions may function like topographically defined data registers, encoding the fluctuating values of abstract semantic variables. This functional architecture, which in key respects resembles that of a classical computer, may play a critical role in enabling humans to flexibly generate complex thoughts.Yesterday, the world’s tallest woman was serenaded by 30 pink elephants. The previous sentence is false, but perfectly comprehensible, despite the improbability of the situation it describes. It is comprehensible because the human mind can flexibly combine the meanings of individual words (“woman,” “serenade,” “elephants,” etc.) to compose structured thoughts, such as the meaning of the aforementioned sentence (1, 2). How the brain accomplishes this remarkable feat remains a central, but unanswered, question in cognitive science.Given the vast number of sentences we can understand and produce, it would be implausible for the brain to allocate individual neurons to represent each possible sentence meaning. Instead, it is likely that the brain employs a system for flexibly combining representations of simpler meanings to compose more complex meanings. By “flexibly,” we mean that the same meanings can be combined in many different ways to produce many distinct complex meanings. How the brain flexibly composes complex, structured meanings out of simpler ones is a matter of long-standing debate (3–10).At the cognitive level, theorists have held that the mind encodes sentence-level meaning by explicitly representing and updating the values of abstract semantic variables (3, 5) in a manner analogous to that of a classical computer. Such semantic variables correspond to basic, recurring questions of meaning such as “Who did it?” and “To whom was it done?” On such a view, the meaning of a simple sentence is partly represented by filling in these variables with representations of the appropriate semantic components. For example, “the dog bit the man” would be built out of the same semantic components as “the man bit the dog,” but with a reversal in the values of the “agent” variable (“Who did it?”) and the “patient” variable (“To whom was it done?”). Whether and how the human brain does this remains unknown.Previous research has implicated a network of cortical regions in high-level semantic processing. Many of these regions surround the left sylvian fissure (11–19), including regions of the inferior frontal cortex (13, 14), inferior parietal lobe (12, 20), much of the superior temporal sulcus and gyrus (12, 15, 21), and the anterior temporal lobes (17, 20, 22). Here, we describe two functional magnetic resonance imaging (fMRI) experiments aimed at understanding how the brain (in these regions or elsewhere) flexibly encodes the meanings of sentences involving an agent (“Who did it?”), an action (“What was done?”), and a patient (“To whom was it done?”).First, experiment 1 aims to identify regions that encode structure-dependent meaning. Here, we search for regions that differentiate between pairs of visually presented sentences, where these sentences convey different meanings using the same words (as in “man bites dog” and “dog bites man”). Experiment 1 identifies a region of left mid-superior temporal cortex (lmSTC) encoding structure-dependent meaning. Experiment 2 then asks how the lmSTC represents structure-dependent meaning. Specifically, we test the long-standing hypothesis that the brain represents and updates the values of abstract semantic variables (3, 5): here, the agent (“Who did it?”) and the patient (“To whom was it done?”). We search for distinct neural populations in lmSTC that encode these variables, analogous to the data registers of a computer (5).     

Ras inhibitor CAPRI enables neutrophil-like cells to chemotax through a higher-concentration range of gradients

Neutrophils sense and migrate through an enormous range of chemoattractant gradients through adaptation. Here, we reveal that in human neutrophils, calcium-promoted Ras inactivator (CAPRI) locally controls the GPCR-stimulated Ras adaptation. Human neutrophils lacking CAPRI (capri^kd) exhibit chemoattractant-induced, nonadaptive Ras activation; significantly increased phosphorylation of AKT, GSK-3α/3β, and cofilin; and excessive actin polymerization. capri^kd cells display defective chemotaxis in response to high-concentration gradients but exhibit improved chemotaxis in low- or subsensitive-concentration gradients of various chemoattractants, as a result of their enhanced sensitivity. Taken together, our data reveal that CAPRI controls GPCR activation-mediated Ras adaptation and lowers the sensitivity of human neutrophils so that they are able to chemotax through a higher-concentration range of chemoattractant gradients.

Neutrophils provide first-line host defense and play pivotal roles in innate and adaptive immunity (1–3). The inappropriate recruitment and dysregulated activation of neutrophils contribute to tissue damage and cause autoimmune and inflammatory diseases (1, 4). Neutrophils sense chemoattractants and migrate to sites of inflammation using G protein–coupled receptors (GPCRs). To accurately navigate through an enormous concentration–range gradient of various chemoattractants (10⁻⁹ to ∼10⁻⁵ M; SI Appendix, Fig. S1), neutrophils employ a mechanism called adaptation, in which they no longer respond to present stimuli but remain sensitive to stronger stimuli. Homogeneous, sustained chemoattractant stimuli trigger transient, adaptive responses in many steps of the GPCR-mediated signaling pathway downstream of heterotrimeric G proteins (5, 6). Adaptation provides a fundamental strategy for eukaryotic cell chemotaxis through large concentration–range gradients of chemoattractants. Abstract models and computational simulations have proposed mechanisms generating the temporal dynamics of adaptation: An increase in receptor occupancy activates two antagonistic signaling processes, namely, a rapid “excitation” that triggers cellular responses and a temporally delayed “inhibition” that terminates the responses and results in adaptation (5, 7–13). Many excitatory components have been identified during last two decades; however, the inhibitor(s) have just begun to be revealed (11, 14–17). It has been recently shown that an elevated Ras activity increases the sensitivity and changes migration behavior (18, 19). However, the molecular connection between the GPCR-mediated adaptation and the cell sensitivity remains missing.The small GTPase Ras mediates multiple signaling pathways that control directional cell migration in both neutrophils and Dictyostelium discoideum (17, 20–24). In D. discoideum, Ras is the first signal event that displays GPCR-mediated adaptation (20). Ras signaling is mainly regulated through its activator, guanine nucleotide exchange factor (GEF), and its inactivator, GTPase-activating proteins (GAP) (16, 17, 25). In D. discoideum, the roles of DdNF1 and an F-actin–dependent, negative feedback mechanism have been previously reported (14, 17). We have previously demonstrated the involvement of locally recruited inhibitors that act on upstream of PI₃K in the sensing of chemoattractant gradients (11, 26). Recently, we identified a locally recruited RasGAP protein, C2GAP1, that is essential for F-actin–independent Ras adaptation and long-range chemotaxis in Dictyostelium (16). Active Ras proteins enrich at the leading edge in both D. discoideum cells and neutrophils (17, 27, 28). It has been reported that a RasGEF, RasGRP4, plays a critical role in Ras activation in murine neutrophil chemotaxis (21, 29). However, the components involved in the GPCR-mediated deactivation of Ras and their function in neutrophil chemotaxis are still not known.In the present study, we show that a calcium-promoted Ras inactivator (CAPRI) locally controls the GPCR-mediated Ras adaptation in human neutrophils. In response to high-concentration stimuli, cells lacking CAPRI (capri^kd) exhibit nonadaptive Ras activation; significantly increased activation of AKT, GSK-3α/3β, and cofilin; excessive actin polymerization; and subsequent defective chemotaxis. Unexpectedly, capri^kd cells display enhanced sensitivity toward chemoattractants and an improved chemotaxis in low- or subsensitive-concentration gradients. Taken together, our findings show that CAPRI functions as an inhibitory component of Ras signaling, plays a critical role in controlling the concentration range of chemoattractant sensing, and is important for the proper adaptation during chemotaxis.     

A functional division of Drosophila sweet taste neurons that is value-based and task-specific

Sucrose is an attractive feeding substance and a positive reinforcer for Drosophila. But Drosophila females have been shown to robustly reject a sucrose-containing option for egg-laying when given a choice between a plain and a sucrose-containing option in specific contexts. How the sweet taste system of Drosophila promotes context-dependent devaluation of an egg-laying option that contains sucrose, an otherwise highly appetitive tastant, is unknown. Here, we report that devaluation of sweetness/sucrose for egg-laying is executed by a sensory pathway recruited specifically by the sweet neurons on the legs of Drosophila. First, silencing just the leg sweet neurons caused acceptance of the sucrose option in a sucrose versus plain decision, whereas expressing the channelrhodopsin CsChrimson in them caused rejection of a plain option that was “baited” with light over another that was not. Analogous bidirectional manipulations of other sweet neurons did not produce these effects. Second, circuit tracing revealed that the leg sweet neurons receive different presynaptic neuromodulations compared to some other sweet neurons and were the only ones with postsynaptic partners that projected prominently to the superior lateral protocerebrum (SLP) in the brain. Third, silencing one specific SLP-projecting postsynaptic partner of the leg sweet neurons reduced sucrose rejection, whereas expressing CsChrimson in it promoted rejection of a light-baited option during egg-laying. These results uncover that the Drosophila sweet taste system exhibits a functional division that is value-based and task-specific, challenging the conventional view that the system adheres to a simple labeled-line coding scheme.

The taste systems of many animal species are known to possess a dedicated “channel” for detecting sugars, a class of chemicals that is highly nutritious. For example, mice have been shown to encode gustatory receptors that specifically sense sugars, and the taste neurons that express these sugar receptors on their tongues generally do not express receptors that sense chemicals of another taste modality (e.g., bitterness) (1–3). Furthermore, activation of these sugar-sensing taste neurons by artificial means has been shown to be able to drive appetitive sugar-induced innate responses (e.g., licking) and act as a positive reinforcer for learning (3–5). In some recent studies, these properties of the sweet taste neurons have been found to be present in some of their central nervous system (CNS) targets (e.g., taste-sensitive neurons in the insular cortex), too (6, 7). Thus, one school of thought is that taste coding for sweetness in mice may follow the simple “labeled-line” rule: sweet taste neurons, and potentially some of their central targets, are hardwired to detect sugars specifically and drive sugar-induced reinforcing neural signals and appetitive behaviors (1–7).Drosophila melanogaster also possess sugar-detecting taste neurons. Pioneering early studies have shown that sugar-sensing taste neurons in flies are molecularly, anatomically, and functionally distinct from taste neurons that sense bitterness; sweet-sensing and bitter-sensing taste neurons express different gustatory receptors, project axons to different areas in the brain, and are required to promote different (appetitive versus aversive) behaviors (8–12). Moreover, the activation of sweet neurons by artificial means can drive appetitive behaviors and act as a positive reinforcer for learning (10, 13, 14), while artificial activation of bitter-sensing neurons can induce rejection behaviors and be used as a punishment for learning (10, 13, 15). Interestingly, while these results suggest that Drosophila sweet neurons and their mammalian counterparts have some shared properties, subsequent studies suggest that significant differences exist between them, too. First, the Drosophila genome appears to encode many more sweet receptors than mouse genome does (12, 16–23). Second, Drosophila sweet neurons appear to be able to detect some chemicals that belong to another taste modality [e.g., acetic acid (AA)] (24–27). Third, Drosophila sweet neurons can be found on several body parts (e.g., proboscis and legs) (8, 12, 18, 20, 23, 28–30). Interestingly, sweet neurons on different body parts of Drosophila do not promote identical behavioral outputs (8, 20, 23, 24, 28, 29). For example, labellar sweet neurons and esophageal sweet neurons on the proboscis have been shown to promote proboscis extension reflex (PER) and ingestion, respectively, whereas leg sweet neurons have been shown to promote PER and slowing down of locomotion (8, 12, 28, 29). Collectively, these results suggest that in contrast to the apparent homogeneity of sweet neurons in some mammals, a functional division exists among Drosophila sweet neurons, although the different behavioral responses promoted by different Drosophila sweet neurons generally appear appetitive in nature.In this work, we report yet another striking feature of Drosophila sweet neurons that sets them apart from their mammalian counterparts, namely a functional division that is value-based and task-specific. We discovered this by taking advantage of a context-dependent but highly robust sugar rejection behavior exhibited by egg-laying females (31–34). Previous studies have shown that when selecting for egg-laying site in a small enclosure (dimension ∼16 × 10 × 18 mm), Drosophila readily accept a sucrose-containing agarose for egg-laying when it is the sole option but strongly reject it when a plain option is also available (31, 32). Importantly, silencing their sweet neurons causes the females to no longer reject the sucrose option when choosing between the sucrose versus plain options (31, 32). Thus, in addition to promoting appetitive behaviors and acting as a positive reinforcer, activation of sweet neurons on an egg-laying option can also decrease the value of such an option (thereby causing its rejection over an option that does not activate sweet neurons). These observations not only suggest the existence of an apparent “antiappetitive” role of Drosophila sweet neurons when the task of animals is to select for egg-laying sites but also raise a key question as to whether such counterintuitive, value-decreasing property of sweetness detection during egg-laying may be 1) solely an emergent property of specific neurons in the brain that respond similarly to all peripheral sweet neurons but are sensitive to animals’ behavioral goal and context or 2) carried out by specific sweet neurons at the periphery and then transmitted into the brain via a unique neural pathway activated by these neurons. To disambiguate between these possibilities, we genetically targeted different subsets of sweet neurons to assess their circuit properties as well as their behavioral roles as the animals decided in either a regular or a virtual sweet versus plain decision during egg-laying, taking advantage of a high-throughput closed-loop optogenetic stimulation platform we developed recently. Our collective results support the second scenario and suggest that the value-decreasing property of sweetness/sucrose is conveyed specifically by the sweet neurons on the legs of Drosophila—and not by other sweet neurons—and the unique postsynaptic target(s) of the leg sweet neurons that send long-range projections to the superior lateral protocerebrum (SLP) in the brain. These results reveal a previously unappreciated functional and anatomical division of the Drosophila sweet taste neurons that is both task-specific and value-based, pointing to a level of complexity and sophistication that seems unmatched by their mammalian counterparts so far.