From birth to bite: the evolutionary ecology of India's medically most important snake venoms.

BACKGROUND: Snake venoms can exhibit remarkable inter- and intraspecific variation. While diverse ecological and environmental factors are theorised to explain this variation, only a handful of studies have attempted to unravel their precise roles. This knowledge gap not only impedes our understanding of venom evolution but may also have dire consequences on snakebite treatment. To address this shortcoming, we investigated the evolutionary ecology of venoms of Russell's viper (Daboia russelii) and spectacled cobra (Naja naja), India's two clinically most important snakes responsible for an alarming number of human deaths and disabilities. METHODOLOGY: Several individuals (n = 226) of D. russelii and N. naja belonging to multiple clutches (n = 9) and their mothers were maintained in captivity to source ontogenetic stage-specific venoms. Using various in vitro and in vivo assays, we assessed the significance of prey, ontogeny and sex in driving venom composition, function, and potency. RESULTS: Considerable ontogenetic shifts in venom profiles were observed in D. russelii, with the venoms of newborns being many times as potent as juveniles and adults against mammalian (2.3-2.5 ×) and reptilian (2-10 ×) prey. This is the first documentation of the ontogenetic shift in viperine snakes. In stark contrast, N. naja, which shares a biogeographic distribution similar to D. russelii, deployed identical biochemical cocktails across development. Furthermore, the binding kinetics of cobra venom toxins against synthetic target receptors from various prey and predators shed light on the evolutionary arms race. CONCLUSIONS: Our findings, therefore, provide fascinating insights into the roles of ecology and life history traits in shaping snake venoms.

Reproductive biology, length-weight relationship and diet of co-occurring butterfly rays, Gymnura poecilura and Gymnura zonura, in Malaysian waters.

Recent IUCN assessments had resulted in up listing of the status of butterfly rays due to concerns of overfishing, but inadequate biological understanding of these rays prevents meaningful conservation and management measures. Therefore, this study was undertaken to address knowledge gaps in the reproductive biology and diet of longtail butterfly ray (Gymnura poecilura) and zone tail butterfly ray (Gymnura zonura) in Malaysian waters. From surveys of landing sites and fish markets from years 2017 to 2022, size (disc width, DW), weight and maturity were recorded, and stomachs were collected from 94 G. poecilura (N = 39 females and 55 males) and 20 G. zonura (N = 10 females and 10 males) specimens. The length-weight relationships were significantly different between sexes for G. poecilura. The size at maturity (DW50) was estimated to be 476.0 mm (females), 385.0 mm (males) for G. poecilura and 442.0 mm (combined) for G. zonura. The number of embryos ranged from 1 to 6, and the embryo size was between 73.90 to 130.44 mm DW. Dietary analysis of stomach contents revealed that fish prey was dominant in both G. poecilura [94.4% Index of Relative Importance (IRI)] and G. zonura (100% IRI). Ontogenetic shift was seen in G. poecilura that fed on more variety of prey items, including shrimps, squids and crabs with an increase in body size. Both species co-occur all along coastal Malaysia although G. zonura is rarely encountered from fisheries surveys along the Strait of Malacca. Given similar habitat associations and dietary habits, G. poecilura may be able to outcompete G. zonura across their shared habitat range. The validity of G. japonica and G. micrura records in Malaysia remains questionable and requires future investigation.

Space-for-time inferences about range-edge dynamics of tree species can be influenced by sampling biases.

Differences between the distributions of tree saplings and adults in geographic or niche space have been used to infer climate change effects on tree range dynamics. Previous studies have reported narrower latitudinal or climatic niche ranges of juvenile trees compared to adults, concluding that tree ranges are contracting, contradicting climate-based predictions. However, more comprehensive sampling of adult trees than juvenile trees in most regional forest inventories could potentially bias ontogenetic comparisons. Here we first report spatial simulations showing that reduced sampling intensity can result in underestimates of range and niche limits, but that resampling the same number of individuals of different life stages can eliminate this bias. We then reanalyzed the U.S. Forest Inventory and Analysis data, comparing the range and niche limits between adult trees and saplings of 92 tree species, both using the original data and two resampling procedures. Resampling aimed to reduce sampling biases by controlling for either sampling area or the number of individuals sampled. Overall, these resampling procedures had a major influence on the estimation of range limits, most often by reducing, eliminating, or even reversing the tendency in the original analyses for saplings to have broader distributions than adult trees. These results indicate that previous conclusions that the distributions of juvenile trees were contracting in response to climate change were potentially artifacts of sampling in the underlying data. More generally, sampling effects involved in the estimation of geographic ranges and environmental niche widths need to be taken into account in studies comparing different life stages, and also likely in other types of distribution comparisons.

Shrub facilitation of tree establishment varies with ontogenetic stage across environmental gradients.

Plant-plant interactions are important drivers of ecosystem structure and function, yet predicting interaction outcomes across environmental gradients remains challenging. Understanding how interactions are affected by ontogenetic shifts in plant characteristics can provide insight into the drivers of interactions and improve our ability to anticipate ecosystem responses to environmental change. We developed a conceptual framework of nurse shrub facilitation of tree establishment. We used a combination of field experiments and environmental measurements to test the framework with a shrub (Artemisia tridentata) and a tree (Pinus monophylla), two foundation species in a semiarid environment. Shrub microsites allowed trees to overcome an early population bottleneck and successfully establish in areas without tree cover. Shrubs facilitated trees at multiple ontogenetic stages, but the net outcome of the interaction shifted from strongly positive to neutral after the transition of P. monophylla from juvenile to adult foliage. Microhabitat conditions varied across a broad elevational gradient, but interaction outcomes were not strongly related to elevation. Favorable microsites provided by A. tridentata cover are crucial for P. monophylla recovery after stand-replacing disturbance. Models of vegetation response to rapid global environmental change should incorporate the critically important role of nurse shrub interactions for ameliorating population bottlenecks in tree establishment.

Lack of social preference between unfamiliar and familiar juvenile Port Jackson sharks Heterodontus portusjacksoni.

This study investigated whether captive-reared juvenile Port Jackson sharks Heterodontus portusjacksoni choose to aggregate and if familiarity is one of the mechanisms driving social preference. In a controlled binary-choice experiment, juvenile sharks were given the option to associate or not with unfamiliar conspecifics, or to associate or not with familiar conspecifics. In neither group did juvenile H. portusjacksoni actively choose to associate with conspecifics, but familiarity decreased the proportion of time spent near a conspecific only during the initial phase of the experiment. Treatment (1 or 3 shoal mates), sex and size had no effect on aggregation behaviour. These findings suggest that familiarity is not a driver of social preferences in juvenile H. portusjacksoni, contrary to results in another shark species. Additionally, adult H. portusjacksoni form large aggregations during the breeding season and actively associate with familiar sex and size-matched individuals, thus our results suggest the species undergoes an ontogenetic shift in social behaviour.

Phenological mismatch and ontogenetic diet shifts interactively affect offspring condition in a passerine.

Climate change may cause phenological asynchrony between trophic levels, which can lead to mismatched reproduction in animals. Although indirect effects of mismatch on fitness are well described, direct effects on parental prey choice are not. Moreover, direct effects of prey variation on offspring condition throughout their early development are understudied. Here, we used camera trap data collected over 2 years to study the effects of trophic mismatch and nestling age on prey choice in pied flycatchers (Ficedula hypoleuca). Furthermore, we studied the effect of mismatch and variation in nestling diet on offspring condition. Both experimentally induced and natural mismatches with the caterpillar peak negatively affected absolute and relative numbers of caterpillars and offspring condition (mass, tarsus and wing length) and positively affected absolute and relative numbers of flying insects in the nestling diet. Feeding more flying insects was negatively correlated with nestling day 12 mass. Both descriptive and experimental data showed preferential feeding of spiders when nestlings were <7 days old. Receiving more spiders during this phase was positively correlated with tarsus growth. These results highlight the need for a more inclusive framework to study phenological mismatch in nature. The general focus on only one prey type, the rarity of studies that measure environmental abundance of prey, and the lack of timing experiments in dietary studies currently hamper understanding of the actual trophic interactions that affect fitness under climate change.

Explaining ontogenetic shifts in root-shoot scaling with transient dynamics.

BACKGROUND AND AIMS: Simple models of herbaceous plant growth based on optimal partitioning theory predict, at steady state, an isometric relationship between shoot and root biomass during plant ontogeny, i.e. a constant root-shoot ratio. This prediction has received mixed empirical support, suggesting either that optimal partitioning is too coarse an assumption to model plant biomass allocation, or that additional processes need to be modelled to account for empirical findings within the optimal partitioning framework. In this study, simulations are used to compare quantitatively two potential explanations for observed non-isometric relationships, namely nutrient limitation during the experiments and initial developmental constraints. METHODS: A simple plant growth model was built to simulate the growth of herbaceous species, based on optimal partitioning theory combined with empirically measured plant functional traits. Its ability to reproduce plant relative growth rate and final root weight ratio was assessed against previously published data. Predicted root-shoot ratios during plant ontogeny were compared with experimental observations. The effects of nutrient limitation and initial developmental constraints on root-shoot ratios were then tested. KEY RESULTS: The model was found to reproduce overall plant growth patterns accurately, but failed, in its simplest form, at explaining non-isometric growth trajectories. Both nutrient limitation and ontogenetic developmental constraints were further shown to cause transient dynamics resulting in a deviation from isometry. Nitrogen limitation alone was not sufficient to explain the observed trajectories of most plant species. The inclusion of initial developmental constraints (fixed non-optimal initial root-shoot ratios) enabled the reproduction of the observed trajectories and were consistent with observed initial root-shoot ratios. CONCLUSIONS: This study highlights the fact that considering transient dynamics enables theoretical predictions based on optimal partitioning to be reconciled with empirically measured ontogenetic root-shoot allometries. The transient dynamics cannot be solely explained by nutrient limitation during the experiments, pointing to a likely role for initial developmental constraints in the observed non-isometric growth trajectories.

Fine-scale foraging ecology and habitat use of sympatric green and hawksbill turtles in the Western Indian ocean.

Using stable isotope analysis of carbon and nitrogen of turtle tissues and putative prey items, we investigated the diet of immature green turtles and hawksbill turtles foraging in the lagoon of Aldabra Atoll, a relatively undisturbed atoll in the southern Seychelles. Aldabra offers a unique environment for understanding sea turtle ecology. Green turtles mostly consumed seagrass and brown algae while hawksbill turtles mainly consumed mangroves and invertebrates. Green turtles showed a dietary shift with size (a proxy for age). There was minimal niche overlap between species and evidence of small-scale foraging site fidelity with turtle tissue reflecting site-specific prey. This highlights the ecological importance of seagrass and mangrove habitats and suggests that turtles play a role in controlling algal biomass at Aldabra. This study is the first to closely examine the foraging ecology of these sympatric turtle species in the Western Indian Ocean, a globally important region for both species.

Ontogenetic shifts in plant interactions vary with environmental severity and affect population structure.

Environmental conditions and plant size may both alter the outcome of inter-specific plant-plant interactions, with seedlings generally facilitated more strongly than larger individuals in stressful habitats. However, the combined impact of plant size and environmental severity on interactions is poorly understood. Here, we tested explicitly for the first time the hypothesis that ontogenetic shifts in interactions are delayed under increasingly severe conditions by examining the interaction between a grass, Agrostis magellanica, and a cushion plant, Azorella selago, along two severity gradients. The impact of A. selago on A. magellanica abundance, but not reproductive effort, was related to A. magellanica size, with a trend for delayed shifts towards more negative interactions under greater environmental severity. Intermediate-sized individuals were most strongly facilitated, leading to differences in the size-class distribution of A. magellanica on the soil and on A. selago. The A. magellanica size-class distribution was more strongly affected by A. selago than by environmental severity, demonstrating that the plant-plant interaction impacts A. magellanica population structure more strongly than habitat conditions. As ontogenetic shifts in plant-plant interactions cannot be assumed to be constant across severity gradients and may impact species population structure, studies examining the outcome of interactions need to consider the potential for size- or age-related variation in competition and facilitation.

Ontogenetic resource use and trophic dynamics of endangered juvenile Tachypleus tridentatus among diversified nursery habitats in the northern Beibu Gulf, China.

Horseshoe crabs, the most well-known example of "living fossils", are iconic and ecologically important macroinvertebrates in coastal and estuarine ecosystems. Their blood is a crucial resource for manufacturing Limulus or Tachypleus amebocyte lysate to detect bacterial endotoxins or fungal contamination in drug and medical devices. An enhanced understanding of their ecological roles and trophic interactions in the food webs is fundamental to facilitate resource management for the declining populations in Asia. Foraging information of the Asian species, however, is mainly derived from preliminary, scattered reports from a limited number of study locations. In this study, resource utilization, trophic niche dynamics, and trophic interaction of the juvenile tri-spine horseshoe crab, Tachypleus tridentatus (instars 1-12, approximately 0.5-8 years old) across ontogeny was assessed in diversified nursery habitats along the northern Beibu Gulf, China, using carbon and nitrogen stable isotopes. Our results suggest that: (i) T. tridentatus are ecological generalists given the vast range of carbon isotopic values and trophic niche width estimates exhibited between multiple instar groups; (ii) juvenile T. tridentatus across most habitat types predominantly assimilated energy from a variety of basal production sources in the food web, but primarily depended on sedimentary organic matter and seagrass resource pools; (iii) ontogenetic shifts in juvenile dietary proportions were evident, with decreased reliance on sedimentary organic matter, coupled with increased reliance on benthic macroinvertebrate grazers, detritivores, and omnivores with age; and (iv) nearly all juvenile instars occupied similar trophic positions in the food web with slight shifts in trophic position present with increasing size. Our findings indicate that resource availability and ontogenetic diet shifts strongly influence horseshoe crab trophic dynamics, and age should be accounted when formulating habitat conservation measures based on resource use for Asian horseshoe crabs.

A modified niche model for generating food webs with stage-structured consumers: The stabilizing effects of life-history stages on complex food webs.

Almost all organisms grow in size during their lifetime and switch diets, trophic positions, and interacting partners as they grow. Such ontogenetic development introduces life-history stages and flows of biomass between the stages through growth and reproduction. However, current research on complex food webs rarely considers life-history stages. The few previously proposed methods do not take full advantage of the existing food web structural models that can produce realistic food web topologies.We extended the niche model developed by Williams and Martinez (Nature, 2000, 404, 180-183) to generate food webs that included trophic species with a life-history stage structure. Our method aggregated trophic species based on niche overlap to form a life-history structured population; therefore, it largely preserved the topological structure of food webs generated by the niche model. We applied the theory of allometric predator-prey body mass ratio and parameterized an allometric bioenergetic model augmented with biomass flow between stages via growth and reproduction to study the effects of a stage structure on the stability of food webs.When life-history stages were linked via growth and reproduction, more food webs persisted, and persisting food webs tended to retain more trophic species. Topological differences between persisting linked and unlinked food webs were small to modest. The slopes of biomass spectra were lower, and weak interaction links were more prevalent in the linked food webs than the unlinked ones, suggesting that a life-history stage structure promotes characteristics that can enhance stability of complex food webs.Our results suggest a positive relationship between the complexity and stability of complex food webs. A life-history stage structure in food webs may play important roles in dynamics of and diversity in food webs.

Feeding ecology and microplastic ingestion in Chelon richardsonii (Mugilidae) associated with surf diatom Anaulus australis accumulations in a warm temperate South African surf zone.

The study aimed to explore ontogenetic dietary changes and microplastic ingestion in Chelon richardsonii associated with diatom Anaulus australis accumulations in a warm temperate South African surf zone. This species is commercially important and forms an important trophic link in southern African coastal waters. Postflexion larvae, juveniles and adults are dominant in surf zones and were used for comparison. Index of Relative Importance (%IRI) results showed an ontogenetic dietary shift, with postflexion larvae ingesting mostly copepods (%IRI = 52.0) and microplastic fibres (%IRI = 30.7); while older developmental stages fed predominantly on diatoms (%IRI ranging from 53.9 to 65.6) and sand (%IRI ranging from 34.2 to 46.0). Microplastic fibres were found in 40% of fish sampled, microplastic fragments in 5%, and both types were recorded across all developmental stages assessed. This study provides the first record of microplastics in C. richardsonii, adding to growing microplastic research in fishes.

Beyond polyphagy and opportunism: natural prey of hunting spiders in the canopy of apple trees.

Spiders (Araneae) form abundant and diverse assemblages in agroecosystems such as fruit orchards, and thus might have an important role as natural enemies of orchard pests. Although spiders are polyphagous and opportunistic predators in general, limited information exists on their natural prey at both species and community levels. Thus, the aim of this study was to assess the natural prey (realized trophic niche) of arboreal hunting spiders, their role in trophic webs and their biological control potential with direct observation of predation events in apple orchards. Hunting spiders with prey in their chelicerae were collected in the canopy of apple trees in organic apple orchards in Hungary during the growing seasons between 2013 and 2019 and both spiders and their prey were identified and measured. Among others, the composition of the actual (captured by spiders) and the potential (available in the canopy) prey was compared, trophic niche and food web metrics were calculated, and some morphological, dimensional data of the spider-prey pairs were analyzed. Species-specific differences in prey composition or pest control ability were also discussed. By analyzing a total of 878 prey items captured by spiders, we concluded that arboreal hunting spiders forage selectively and consume a large number of apple pests; however, spiders' beneficial effects are greatly reduced by their high levels of intraguild predation and by a propensity to switch from pests to alternative prey. In this study, arboreal hunting spiders showed negative selectivity for pests, no selectivity for natural enemies and positive selectivity for neutral species. In the trophic web, the dominant hunting spider taxa/groups (Carrhotus xanthogramma, Philodromus cespitum, Clubiona spp., Ebrechtella tricuspidata, Xysticus spp. and 'Other salticids') exhibit different levels of predation on different prey groups and the trophic web's structure changes depending on the time of year. Hunting spiders show a high functional redundancy in their predation, but contrary to their polyphagous nature, the examined spider taxa showed differences in their natural diet, exhibited a certain degree of prey specialization and selected prey by size and taxonomic identity. Guilds (such as stalkers, ambushers and foliage runners) did not consistently predict either prey composition or predation selectivity of arboreal hunting spider species. From the economic standpoint, Ph. cespitum and Clubiona spp. were found to be the most effective natural enemies of apple pests, especially of aphids. Finally, the trophic niche width of C. xanthogramma and Ph. cespitum increased during ontogeny, resulting in a shift in their predation. These results demonstrate how specific generalist predators can differ from each other in aspects of their predation ecology even within a relatively narrow taxonomic group.

A dominance shift in arid savanna: An herbaceous legume outcompetes local C4 grasses.

The characteristic vegetation structure of arid savannas with a dominant layer of perennial grass is maintained by the putative competitive superiority of the C4 grasses. When this competitive balance is disturbed by weakening the grasses or favoring the recruitment of other species, trees, shrubs, single grass, or forb species can increase and initiate sudden dominance shifts. Such shifts involving woody species, often termed "shrub encroachment", or the mass spreading of so-called increaser species have been extensively researched, but studies on similar processes without obvious preceding disturbance are rare. In Namibia, the native herbaceous legume Crotalaria podocarpa has recently encroached parts of the escarpment region, seriously affecting the productivity of local fodder grasses. Here, we studied the interaction between seedlings of the legume and the dominant local fodder grass (Stipagrostis ciliata). We used a pot experiment to test seedling survival and to investigate the growth of Crotalaria in competition with Stipagrostis. Additional field observations were conducted to quantify the interactive effect. We found germination and growth of the legume seedlings to be facilitated by inactive (dead or dormant) grass tussocks and unhindered by active ones. Seedling survival was three times higher in inactive tussocks and Crotalaria grew taller. In the field, high densities of the legume had a clear negative effect on productivity of the grass. The C4 grass was unable to limit the recruitment and spread of the legume, and Crotalaria did outcompete the putative more competitive grass. Hence, the legume is able to spread and establish itself in large numbers and initiate a dominance shift in savannas, similar to shrub encroachment.

Transcriptomics-guided bottom-up and top-down venomics of neonate and adult specimens of the arboreal rear-fanged Brown Treesnake, Boiga irregularis, from Guam.

The Brown Treesnake (Boiga irregularis) is an arboreal, nocturnal, rear-fanged venomous snake native to northern and eastern regions of Australia, Papua New Guinea and the Solomon Islands. It was inadvertently introduced onto the island of Guam during the late 1940's to early 1950's, and it has caused massive declines and extirpations of the native bird, lizard, and mammal populations. In the current study, we report the characterization of the venom proteome of an adult and a neonate B. irregularis specimens from Guam by a combination of venom gland transcriptomic and venomic analyses. Venom gland transcriptomic analysis of an adult individual identified toxins belonging to 18 protein families, with three-finger toxin isoforms being the most abundantly expressed transcripts, comprising 94% of all venom protein transcript reads. Transcripts for PIII-metalloproteinases, C-type lectins, cysteine-rich secretory proteins, acetylcholinesterases, natriuretic peptides, ficolins, phospholipase A2 (PLA2) inhibitors, PLA2s, vascular endothelial growth factors, Kunitz-type protease inhibitors, cystatins, phospholipase Bs, cobra venom factors, waprins, SVMP inhibitors, matrix metalloproteinases, and hyaluronidases were also identified, albeit, at very low abundances ranging from 0.05% to 1.7% of the transcriptome. The venom proteomes of neonate and adult B. irregularis were also both overwhelmingly (78 and 84%, respectively) dominated by monomeric and dimeric 3FTxs, followed by moderately abundant (21% (N) and 13% (A)) CRISPs, low abundance (1% (N), 3% (A)) PIII-SVMPs, and very low abundance (<0.01%) PLA2 and SVMP inhibitors. The differences in relative toxin abundances identified between neonate and adult snakes likely correlates to shifts in prey preference between the two age classes, from nearly-exclusively lizards to lizards, birds and small mammals. Immunoaffinity antivenomics with experimentally designed rabbit anti-Brown Treesnake (anti-BTS) venom IgGs against homologous venom from adult snakes demonstrated that CRISPs, PIII-SVMPs, and 60-70% of 3FTxs were effectively immunocaptured. Western blot analysis showed that all venom proteins were recognized by anti-BTS IgGs, and cross-reactivity with other rear-fanged snake venoms was also observed. Incubation of anti-BTS venom IgGs with crude B. irregularis venom resulted in a significant decrease in proteolytic (SVMP) activity against azocasein. These results provide the first comparative venomic and anti-venomic analysis of neonate and adult B. irregularis from Guam, further highlighting evolutionary trends in venom composition among rear-fanged venomous snakes. SIGNIFICANCE PARAGRAPH: The Brown Treesnake (Boiga irregularis) has caused extensive ecological and economic damage to the island of Guam where it has become a classic example of the negative impacts of invasive species. In the current study, we report the first combined transcriptomic and proteomic analysis of B. irregularis venom of Guam origin. The transcriptome of an adult snake contained toxin sequences belonging to 18 protein families, with three-finger toxin (3FTx) isoforms being the most abundant and representing 94% of all venom protein transcript reads. Our bottom-up and top-down venomic analyses confirmed that 3FTxs are the major components of B. irregularis venom, and a comparative analysis of neonate and adult venoms demonstrate a clear ontogenetic shift in toxin abundance, likely driven by dietary variation between the two age classes. Second-generation antivenomics and Western blot analysis using purified anti-Brown Treesnake rabbit serum IgGs (anti-BTS IgGs) showed strong immunoreactivity toward B. irregularis venom. Interestingly, our anti-BTS IgGs did not cross-react with 3FTxs found in several other rear-fanged snake venoms, or against 3FTxs in the venom of the elapid Ophiophagus hannah, indicating that epitopes in these 3FTx molecules are quite distinct.

The Feeding Ecology of the Blue Swimming Crab, Portunus pelagicus (Linnaeus, 1758), at Kung Krabaen Bay, Chanthaburi Province, Thailand.

The natural diet of blue swimming crabs, Portunus pelagicus, was investigated from October 2008 to October 2009 using hand sampling and a crab gill net. The results showed that the major prey items in the stomach contents of P. pelagicus were teleost fish (29.61%), organic matter (20.69%), crustaceans (18.3%) and shelled molluscs (11.46%). Significant differences were found in diet composition between juvenile and mature crabs, between crabs inside and outside the bay and among seasons. In contrast, significant differences were not found between male and female crabs.