Rapid quantification of biological nitrogen fixation using optical spectroscopy.

Biological nitrogen fixation (BNF) provides a globally important input of nitrogen (N); its quantification is critical but technically challenging. Leaf reflectance spectroscopy offers a more rapid approach than traditional techniques to measure plant N concentration ([N]) and isotopes (δ15N). Here we present a novel method for rapidly and inexpensively quantifying BNF using optical spectroscopy. We measured plant [N], δ15N, and the amount of N derived from atmospheric fixation (Ndfa) following the standard traditional methodology using isotope ratio mass spectrometry (IRMS) from tissues grown under controlled conditions and taken from field experiments. Using the same tissues, we predicted the same three parameters using optical spectroscopy. By comparing the optical spectroscopy-derived results with traditional measurements (i.e. IRMS), the amount of Ndfa predicted by optical spectroscopy was highly comparable to IRMS-based quantification, with R2 being 0.90 (slope=0.90) and 0.94 (slope=1.02) (root mean square error for predicting legume δ15N was 0.38 and 0.43) for legumes grown in glasshouse and field, respectively. This novel application of optical spectroscopy facilitates BNF studies because it is rapid, scalable, low cost, and complementary to existing technologies. Moreover, the proposed method successfully captures the dynamic response of BNF to climate changes such as warming and drought.

Warmer ambient temperatures reduce protein intake by a mammalian folivore.

The interplay between ambient temperature and nutrition in wild herbivores is frequently overlooked, despite the fundamental importance of food. We tested whether different ambient temperatures (10°C, 18°C and 26°C) influenced the intake of protein by a marsupial herbivore, the common brushtail possum (Trichosurus vulpecula). At each temperature, possums were offered a choice of two foods containing different amounts of protein (57% versus 8%) for one week. Animals mixed a diet with a lower proportion of protein to non-protein (P : NP, 0.20) when held at 26°C compared to that at both 10°C and 18°C (0.22). Since detoxification of plant secondary metabolites imposes a protein cost on animals, we then studied whether addition of the monoterpene 1,8-cineole to the food changed the effect of ambient temperature (10°C and 26°C) on food choice. Cineole reduced food intake but also removed the effect of temperature on P : NP ratio and instead animals opted for a diet with higher P : NP (0.19 with cineole versus 0.15 without cineole). These experiments show the proportion of P : NP chosen by animals is influenced by ambient temperature and by plant secondary metabolites. Protein is critical for reproductive success in this species and reduced protein intake caused by high ambient temperatures may limit the viability of some populations in the future. This article is part of the theme issue 'Food processing and nutritional assimilation in animals'.

The koala gut microbiome is largely unaffected by host translocation but rather influences host diet.

Introduction: Translocation is a valuable and increasingly used strategy for the management of both threatened and overabundant wildlife populations. However, in some instances the translocated animals fail to thrive. Differences in diet between the source and destination areas may contribute to poor translocation outcomes, which could conceivably be exacerbated if the animals' microbiomes are unsuited to the new diet and cannot adapt. Methods: In this study we tracked how the faecal microbiome of a specialist Eucalyptus folivore, the koala (Phascolarctos cinereus), changed over the course of a year after translocation. We assessed microbiome composition by 16S rRNA amplicon sequencing of faecal pellets. Results: We found no significant overall changes in the faecal microbiomes of koalas post-translocation (n = 17) in terms of microbial richness, diversity or composition when compared to the faecal microbiomes of koalas from an untranslocated control group (n = 12). This was despite the translocated koalas feeding on a greater variety of Eucalyptus species after translocation. Furthermore, while differences between koalas accounted for half of the microbiome variation, estimated diets at the time of sampling only accounted for 5% of the variation in the koala microbiomes between sampling periods. By contrast, we observed that the composition of koala faecal microbiomes at the time of translocation accounted for 37% of between koala variation in post-translocation diet. We also observed that translocated koalas lost body condition during the first month post-translocation and that the composition of the koalas' initial microbiomes were associated with the magnitude of that change. Discussion: These findings suggest that the koala gut microbiome was largely unaffected by dietary change and support previous findings suggesting that the koala gut microbiome influences the tree species chosen for feeding. They further indicate that future research is needed to establish whether the koalas' gut microbiomes are directly influencing their health and condition or whether aspects of the koala gut microbiomes are an indicator of underlying physiological differences or pathologies. Our study provides insights into how animal microbiomes may not always be affected by the extreme upheaval of translocation and highlights that responses may be host species-specific. We also provide recommendations to improve the success of koala translocations in the future.

Individuality and stability of the koala (Phascolarctos cinereus) faecal microbiota through time.

Gut microbiota studies often rely on a single sample taken per individual, representing a snapshot in time. However, we know that gut microbiota composition in many animals exhibits intra-individual variation over the course of days to months. Such temporal variations can be a confounding factor in studies seeking to compare the gut microbiota of different wild populations, or to assess the impact of medical/veterinary interventions. To date, little is known about the variability of the koala (Phascolarctos cinereus) gut microbiota through time. Here, we characterise the gut microbiota from faecal samples collected at eight timepoints over a month for a captive population of South Australian koalas (n individuals = 7), and monthly over 7 months for a wild population of New South Wales koalas (n individuals = 5). Using 16S rRNA gene sequencing, we found that microbial diversity was stable over the course of days to months. Each koala had a distinct faecal microbiota composition which in the captive koalas was stable across days. The wild koalas showed more variation across months, although each individual still maintained a distinct microbial composition. Per koala, an average of 57 (±16) amplicon sequence variants (ASVs) were detected across all time points; these ASVs accounted for an average of 97% (±1.9%) of the faecal microbial community per koala. The koala faecal microbiota exhibits stability over the course of days to months. Such knowledge will be useful for future studies comparing koala populations and developing microbiota interventions for this regionally endangered marsupial.

Patch quality and habitat fragmentation shape the foraging patterns of a specialist folivore.

Research on use of foraging patches has focused on why herbivores visit or quit patches, yet little is known about visits to patches over time. Food quality, as reflected by higher nutritional quality and lower plant defenses, and physical patch characteristics, which offer protection from predators and weather, affect patch use and hence should influence their revisitation. Due to the potentially high costs of moving between patches, fragmented habitats are predicted to complicate foraging decisions of many animals. We aimed to determine how food quality, shelter availability and habitat fragmentation influence tree reuse by a specialist folivore, the koala, in a fragmented agricultural landscape. We GPS-tracked 23 koalas in northern New South Wales, Australia and collated number of revisits, average residence time, and average time-to-return to each tree. We measured tree characteristics including food quality (foliar nitrogen and toxic formylated phloroglucinol compounds, FPCs concentrations), tree size, and tree connectedness. We also modeled the costs of locomotion between trees. Koalas re-visited isolated trees with high leaf nitrogen disproportionately often. They spent longer time in trees with high leaf nitrogen, and in large trees used for shelter. They took longer to return to trees with low leaf nitrogen. Tree connectivity reduced travel costs between patches, being either individual or groups of trees. FPC levels had no detectable effect on patch revisitation. We conclude that food quality and shelter drive koala tree re-visits. Scattered, isolated trees with nutrient-rich leaves are valuable resource patches for koalas despite movement costs to reach them.

Geographic patterns of koala retrovirus genetic diversity, endogenization, and subtype distributions.

Koala retrovirus (KoRV) subtype A (KoRV-A) is currently in transition from exogenous virus to endogenous viral element, providing an ideal system to elucidate retroviral-host coevolution. We characterized KoRV geography using fecal DNA from 192 samples across 20 populations throughout the koala's range. We reveal an abrupt change in KoRV genetics and incidence at the Victoria/New South Wales state border. In northern koalas, pol gene copies were ubiquitously present at above five per cell, consistent with endogenous KoRV. In southern koalas, pol copies were detected in only 25.8% of koalas and always at copy numbers below one, while the env gene was detected in all animals and in a majority at copy numbers above one per cell. These results suggest that southern koalas carry partial endogenous KoRV-like sequences. Deep sequencing of the env hypervariable region revealed three putatively endogenous KoRV-A sequences in northern koalas and a single, distinct sequence present in all southern koalas. Among northern populations, env sequence diversity decreased with distance from the equator, suggesting infectious KoRV-A invaded the koala genome in northern Australia and then spread south. The exogenous KoRV subtypes (B to K), two novel subtypes, and intermediate subtypes were detected in all northern koala populations but were strikingly absent from all southern animals tested. Apart from KoRV subtype D, these exogenous subtypes were generally locally prevalent but geographically restricted, producing KoRV genetic differentiation among northern populations. This suggests that sporadic evolution and local transmission of the exogenous subtypes have occurred within northern Australia, but this has not extended into animals within southern Australia.

Pastures and Climate Extremes: Impacts of Cool Season Warming and Drought on the Productivity of Key Pasture Species in a Field Experiment.

Shifts in the timing, intensity and/or frequency of climate extremes, such as severe drought and heatwaves, can generate sustained shifts in ecosystem function with important ecological and economic impacts for rangelands and managed pastures. The Pastures and Climate Extremes experiment (PACE) in Southeast Australia was designed to investigate the impacts of a severe winter/spring drought (60% rainfall reduction) and, for a subset of species, a factorial combination of drought and elevated temperature (ambient +3°C) on pasture productivity. The experiment included nine common pasture and Australian rangeland species from three plant functional groups (C3 grasses, C4 grasses and legumes) planted in monoculture. Winter/spring drought resulted in productivity declines of 45% on average and up to 74% for the most affected species (Digitaria eriantha) during the 6-month treatment period, with eight of the nine species exhibiting significant yield reductions. Despite considerable variation in species' sensitivity to drought, C4 grasses were more strongly affected by this treatment than C3 grasses or legumes. Warming also had negative effects on cool-season productivity, associated at least partially with exceedance of optimum growth temperatures in spring and indirect effects on soil water content. The combination of winter/spring drought and year-round warming resulted in the greatest yield reductions. We identified responses that were either additive (Festuca), or less-than-additive (Medicago), where warming reduced the magnitude of drought effects. Results from this study highlight the sensitivity of diverse pasture species to increases in winter and spring drought severity similar to those predicted for this region, and that anticipated benefits of cool-season warming are unlikely to be realized. Overall, the substantial negative impacts on productivity suggest that future, warmer, drier climates will result in shortfalls in cool-season forage availability, with profound implications for the livestock industry and natural grazer communities.

Characterization of the juvenile koala gut microbiome across wild populations.

In this study we compared the faecal microbiomes of wild joey koalas (Phascolarctos cinereus) to those of adults, including their mothers, to establish whether gut microbiome maturation and inheritance in the wild is comparable to that seen in captivity. Our findings suggest that joey koala microbiomes slowly shift towards an adult assemblage between 6 and 12 months of age, as the microbiomes of 9-month-old joeys were more similar to those of adults than those of 7-month-olds, but still distinct. At the phylum level, differences between joeys and adults were broadly consistent with those in captivity, with Firmicutes increasing in relative abundance over the joeys' development and Proteobacteria decreasing. Of the fibre-degrading genes that increased in abundance over the development of captive joeys, those involved in hemicellulose and cellulose degradation, but not pectin degradation, were also generally found in higher abundance in adult wild koalas compared to 7-month-olds. Greater maternal inheritance of the faecal microbiome was seen in wild than in captive koalas, presumably due to the more solitary nature of wild koalas. This strong maternal inheritance of the gut microbiome could contribute to the development of localized differences in microbiome composition, population health and diet through spatial clustering of relatives.

Maternal inheritance of the koala gut microbiome and its compositional and functional maturation during juvenile development.

The acquisition and maturation of the gastrointestinal microbiome is a crucial aspect of mammalian development, particularly for specialist herbivores such as the koala (Phascolarctos cinereus). Joey koalas are thought to be inoculated with microorganisms by feeding on specialized maternal faeces (pap). We found that compared to faeces, pap has higher microbial density, higher microbial evenness and a higher proportion of rare taxa, which may facilitate the establishment of those taxa in joey koalas. We show that the microbiomes of captive joey koalas were on average more similar to those of their mothers than to other koalas, indicating strong maternal inheritance of the faecal microbiome, which can lead to intergenerational gut dysbiosis when the mother is ill. Directly after pap feeding, the joey koalas' microbiomes were enriched for milk-associated bacteria including Bacteroides fragilis, suggesting a conserved role for this species across mammalian taxa. The joeys' microbiomes then changed slowly over 5 months to resemble those of adults by 1 year of age. The relative abundance of fibrolytic bacteria and genes involved in the degradation of plant cell walls also increased in the infants over this time, likely in response to an increased proportion of Eucalyptus leaves in their diets.

Fundamental dietary specialisation explains differential use of resources within a koala population.

The diets of individual animals within populations can differ, but few studies determine whether this is due to fundamental differences in preferences or capacities to eat specific foods, or to external influences such as dominance hierarchies or spatial variation in food availability. The distinction is important because different drivers of dietary specialisation are likely to have different impacts on the way in which animal populations respond to, for example, habitat modification. We used a captive feeding study to investigate the mechanisms driving individual dietary specialisation in a population of wild koalas (Phascolarctos cinereus) in which individuals predominantly ate either Eucalyptus viminalis or Eucalyptus obliqua foliage. All six koalas that primarily ate E. viminalis in the wild avoided eating E. obliqua for more than 1 month in captivity. In contrast, all seven koalas that primarily ate E. obliqua could be maintained exclusively on this species in captivity, although they ate less from individual trees with higher foliar concentrations of unsubstituted B-ring flavanones (UBFs). Our results show that fundamental differences between individual animals allow some to exploit food resources that are less suitable for others. This could reduce competition for food, increase habitat carrying capacity, and is also likely to buffer the population against extinction in the face of habitat modification. The occurrence of fundamental individual specialisation within animal populations could also affect the perceived conservation value of different habitats, translocation or reintroduction success, and population dynamics. It should therefore be further investigated in other mammalian herbivore species.

Mapping canopy nitrogen-scapes to assess foraging habitat for a vulnerable arboreal folivore in mixed-species Eucalyptus forests.

Herbivore foraging decisions are closely related to plant nutritional quality. For arboreal folivores with specialized diets, such as the vulnerable greater glider (Petauroides volans), the abundance of suitable forage trees can influence habitat suitability and species occurrence. The ability to model and map foliar nitrogen would therefore enhance our understanding of folivore habitat use at finer scales. We tested whether high-resolution multispectral imagery, collected by a lightweight and low-cost commercial unoccupied aerial vehicle (UAV), could be used to predict total and digestible foliar nitrogen (N and digN) at the tree canopy level and forest stand-scale from leaf-scale chemistry measurements across a gradient of mixed-species Eucalyptus forests in southeastern Australia. We surveyed temperate Eucalyptus forests across an elevational and topographic gradient from sea level to high elevation (50-1200 m a.s.l.) for forest structure, leaf chemistry, and greater glider occurrence. Using measures of multispectral leaf reflectance and spectral indices, we estimated N and digN and mapped N and favorable feeding habitat using machine learning algorithms. Our surveys covered 17 Eucalyptus species ranging in foliar N from 0.63% to 1.92% dry matter (DM) and digN from 0.45% to 1.73% DM. Both multispectral leaf reflectance and spectral indices were strong predictors for N and digN in model cross-validation. At the tree level, 79% of variability between observed and predicted measures of nitrogen was explained. A spatial supervised classification model correctly identified 80% of canopy pixels associated with high N concentrations (≥1% DM). We developed a successful method for estimating foliar nitrogen of a range of temperate Eucalyptus species using UAV multispectral imagery at the tree canopy level and stand scale. The ability to spatially quantify feeding habitat using UAV imagery allows remote assessments of greater glider habitat at a scale relevant to support ground surveys, management, and conservation for the vulnerable greater glider across southeastern Australia.

Silicon Defence in Plants: Does Herbivore Identity Matter?

Silicon accumulation is a key defence against herbivorous pests, but may have wider detrimental impacts if plants become unpalatable for livestock. We argue that some herbivores are better adapted to silicon-rich diets than others; herbivore anatomy and physiology, and the nature of silicon deposition, are crucial to understanding these differences.

The fate of carbon in a mature forest under carbon dioxide enrichment.

Atmospheric carbon dioxide enrichment (eCO2) can enhance plant carbon uptake and growth1-5, thereby providing an important negative feedback to climate change by slowing the rate of increase of the atmospheric CO2 concentration6. Although evidence gathered from young aggrading forests has generally indicated a strong CO2 fertilization effect on biomass growth3-5, it is unclear whether mature forests respond to eCO2 in a similar way. In mature trees and forest stands7-10, photosynthetic uptake has been found to increase under eCO2 without any apparent accompanying growth response, leaving the fate of additional carbon fixed under eCO2 unclear4,5,7-11. Here using data from the first ecosystem-scale Free-Air CO2 Enrichment (FACE) experiment in a mature forest, we constructed a comprehensive ecosystem carbon budget to track the fate of carbon as the forest responded to four years of eCO2 exposure. We show that, although the eCO2 treatment of +150 parts per million (+38 per cent) above ambient levels induced a 12 per cent (+247 grams of carbon per square metre per year) increase in carbon uptake through gross primary production, this additional carbon uptake did not lead to increased carbon sequestration at the ecosystem level. Instead, the majority of the extra carbon was emitted back into the atmosphere via several respiratory fluxes, with increased soil respiration alone accounting for half of the total uptake surplus. Our results call into question the predominant thinking that the capacity of forests to act as carbon sinks will be generally enhanced under eCO2, and challenge the efficacy of climate mitigation strategies that rely on ubiquitous CO2 fertilization as a driver of increased carbon sinks in global forests.

When resistance is futile, tolerate instead: silicon promotes plant compensatory growth when attacked by above- and belowground herbivores.

Plants have evolved numerous herbivore defences that are resistance- or tolerance-based. Resistance involves physical and chemical traits that deter and/or harm herbivores whereas tolerance minimizes fitness costs of herbivory, often via compensatory growth. The Poaceae frequently accumulate large amounts of silicon (Si), which can be used for herbivore resistance, including biomechanical and (indirectly) biochemical defences. To date, it is unclear whether Si improves tolerance of herbivory. Here we report how Si enabled a cereal (Triticum aestivum) to tolerate damage inflicted by above- and belowground herbivores. Leaf herbivory increased Si concentrations in the leaves by greater than 50% relative to herbivore-free plants, indicating it was an inducible defensive response. In plants without Si supplementation, leaf herbivory reduced shoot biomass by 52% and root herbivory reduced root biomass by 68%. Si supplementation, however, facilitated compensatory growth such that shoot losses were more than compensated for (+14% greater than herbivore-free plants) and root losses were minimized to -16%. Si supplementation did not improve plant resistance since Si did not enhance biomechanical resistance (i.e. force of fracture) or reduce leaf consumption and herbivore relative growth rates. We propose that Si-based defence operates in wheat via tolerance either in addition or as an alternative to resistance-based defence.

Ingestion and Absorption of Eucalypt Monoterpenes in the Specialist Feeder, the Koala (Phascolarctos cinereus).

The koala is a specialist feeder with a diet consisting almost exclusively of potentially toxic eucalypt leaves. Monoterpenes, an abundant class of plant secondary metabolites in eucalypts, are highly lipophilic. Chronic absorption and systemic exposure can be anticipated for the koala, causing health effects in various ways when consumed in high amounts, but particularly causing alterations in immune function in this species. Therefore, careful leaf selection, efficient detoxification pathways, and other specialist adaptations are required to protect animals from acute intoxication. This is the first paper providing insight into the systemic exposure of koalas to these compounds. Profiles of six selected major monoterpenes were investigated in the ingesta of deceased koalas from four different regions of NSW and South-East Queensland. Concentrations of the same compounds were measured in lymphoid tissues of deceased koalas and in the blood of live koalas from other regions of NSW. Analytical methods included liquid extraction and solid-phase micro-extraction, followed by gas-chromatography/ mass-spectrometry. Concentrations in the ingesta of individual animals vary remarkably, though the average proportions of individual monoterpenes in the ingesta of animals from the four different regions are highly comparable. Blood concentrations of the selected monoterpenes also varied considerably. The highest blood concentrations were found for 1,8-cineole, up to 971 ng/ml. There was similarity between circulating monoterpene profiles and ingesta profiles. Based on the observed lack of similarity between blood and lymph tissue concentrations, individual monoterpenes either exhibit different affinities for lymphatic tissue compared to blood or their accumulation in blood and lymph tissue differs temporally. In general, blood monoterpene concentrations found in koalas were low compared to those reported in other marsupial eucalypt feeders, but significant concentrations of monoterpenes were detected in all samples analysed. This data on blood and lymphatic tissue monoterpene concentrations builds the fundamental groundwork for future research into the effects of dietary monoterpenes on various biological processes of specialist herbivores and into the significance of these animals' metabolic and behavioural strategies for coping with these compounds. We have shown that the systemic exposure of koalas to potentially anti-inflammatory eucalypt monoterpenes is continuous, and we provide data on physiological concentrations which will allow realistic future studies of the effects of monoterpenes on immune cell function.

The Koala (Phascolarctos cinereus) faecal microbiome differs with diet in a wild population.

BACKGROUND: The diet of the koala (Phascolarctos cinereus) is comprised almost exclusively of foliage from the genus Eucalyptus (family Myrtaceae). Eucalyptus produces a wide variety of potentially toxic plant secondary metabolites which have evolved as chemical defences against herbivory. The koala is classified as an obligate dietary specialist, and although dietary specialisation is rare in mammalian herbivores, it has been found elsewhere to promote a highly-conserved but low-diversity gut microbiome. The gut microbes of dietary specialists have been found sometimes to enhance tolerance of dietary PSMs, facilitating competition-free access to food. Although the koala and its gut microbes have evolved together to utilise a low nutrient, potentially toxic diet, their gut microbiome has not previously been assessed in conjunction with diet quality. Thus, linking the two may provide new insights in to the ability of the koala to extract nutrients and detoxify their potentially toxic diet. METHOD: The 16S rRNA gene was used to characterise the composition and diversity of faecal bacterial communities from a wild koala population (n = 32) comprising individuals that predominately eat either one of two different food species, one the strongly preferred and relatively nutritious species Eucalyptus viminalis, the other comprising the less preferred and less digestible species Eucalyptus obliqua. RESULTS: Alpha diversity indices indicated consistently and significantly lower diversity and richness in koalas eating E. viminalis. Assessment of beta diversity using both weighted and unweighted UniFrac matrices indicated that diet was a strong driver of both microbial community structure, and of microbial presence/absence across the combined koala population and when assessed independently. Further, principal coordinates analysis based on both the weighted and unweighted UniFrac matrices for the combined and separated populations, also revealed a separation linked to diet. During our analysis of the OTU tables we also detected a strong association between microbial community composition and host diet. We found that the phyla Bacteroidetes and Firmicutes were co-dominant in all faecal microbiomes, with Cyanobacteria also co-dominant in some individuals; however, the E. viminalis diet produced communities dominated by the genera Parabacteroides and/or Bacteroides, whereas the E. obliqua-associated diets were dominated by unidentified genera from the family Ruminococcaceae. DISCUSSION: We show that diet differences, even those caused by differential consumption of the foliage of two species from the same plant genus, can profoundly affect the gut microbiome of a specialist folivorous mammal, even amongst individuals in the same population. We identify key microbiota associated with each diet type and predict functions within the microbial community based on 80 previously identified Parabacteroides and Ruminococcaceae genomes.

Faecal inoculations alter the gastrointestinal microbiome and allow dietary expansion in a wild specialist herbivore, the koala.

BACKGROUND: Differences between individuals in their gastrointestinal microbiomes can lead to variation in their ability to persist on particular diets. Koalas are dietary specialists, feeding almost exclusively on Eucalyptus foliage but many individuals will not feed on particular Eucalyptus species that are adequate food for other individuals, even when facing starvation. We undertook a faecal inoculation experiment to test whether a koala's gastrointestinal (GI) microbiome influences their diet. Wild-caught koalas that initially fed on the preferred manna gum (Eucalyptus viminalis) were brought into captivity and orally inoculated with encapsulated material derived from faeces from koalas feeding on either the less preferred messmate (E. obliqua; treatment) or manna gum (control). RESULTS: The gastrointestinal microbiomes of wild koalas feeding primarily on manna gum were distinct from those feeding primarily on messmate. We found that the gastrointestinal microbiomes of koalas were unresponsive to dietary changes because the control koalas' GI microbiomes did not change even when the nocturnal koalas were fed exclusively on messmate overnight. We showed that faecal inoculations can assist the GI microbiomes of koalas to change as the treatment koalas' GI microbiomes became more similar to those of wild koalas feeding on messmate. There was no overall difference between the control and treatment koalas in the quantity of messmate they consumed. However, the greater the change in the koalas' GI microbiomes, the more messmate they consumed after the inoculations had established. CONCLUSIONS: The results suggest that dietary changes can only lead to changes in the GI microbiomes of koalas if the appropriate microbial species are present, and/or that the koala gastrointestinal microbiome influences diet selection.

Silicon uptake by a pasture grass experiencing simulated grazing is greatest under elevated precipitation.

BACKGROUND: Grasses are hyper-accumulators of silicon (Si) and often up-regulate Si following herbivory. Positive correlations exist between Si and plant water content, yet the extent to which Si uptake responses can be mediated by changes in soil water availability has rarely been studied and never, to our knowledge, under field conditions. We used field-based rain-exclusion shelters to investigate how simulated grazing (shoot clipping) and altered rainfall patterns (drought and elevated precipitation, representing 50% and 150% of ambient precipitation levels, respectively) affected initial patterns of root- and shoot-Si uptake in a native Australian grass (Microlaena stipoides) in Si-supplemented and untreated soils. RESULTS: Si supplementation increased soil water retention under ambient and elevated precipitation but not under drought, although this had little effect on Si uptake and growth (tiller numbers or root biomass) of M. stipoides. Changes in rainfall patterns and clipping had strong individual effects on plant growth and Si uptake and storage, whereby clipping increased Si uptake by M. stipoides under all rainfall treatments but to the greatest extent under elevated precipitation. Moreover, above-ground-below-ground Si distribution only changed following elevated precipitation by decreasing the ratio of root:shoot Si concentrations. CONCLUSIONS: Results highlight the importance of soil water availability for Si uptake and suggest a role for both active and passive Si transport mechanisms. Such manipulative field studies may provide a more realistic insight into how grasses initially respond to herbivory in terms of Si-based defence under different environmental conditions.

Spatial correlations between browsing on balsam fir by white-tailed deer and the nutritional value of neighboring winter forage.

Associational effects, that is, the influence of neighboring plants on herbivory suffered by a plant, are an outcome of forage selection. Although forage selection is a hierarchical process, few studies have investigated associational effects at multiple spatial scales. Because the nutritional quality of plants can be spatially structured, it might differently influence associational effects across multiple scales. Our objective was to determine the radius of influence of neighbor density and nutritional quality on balsam fir (Abies balsamea) herbivory by white-tailed deer (Odocoileus virginianus) in winter. We quantified browsing rates on fir and the density and quality of neighboring trees in a series of 10-year-old cutovers on Anticosti Island (Canada). We used cross-correlations to investigate relationships between browsing rates and the density and nutritional quality of neighboring trees at distances up to 1,000 m. Balsam fir and white spruce (Picea glauca) fiber content and dry matter in vitro true digestibility were correlated with fir browsing rate at the finest extra-patch scale (across distance of up to 50 m) and between cutover areas (300-400 m). These correlations suggest associational effects, that is, low nutritional quality of neighbors reduces the likelihood of fir herbivory (associational defense). Our results may indicate associational effects mediated by intraspecific variation in plant quality and suggest that these effects could occur at scales from tens to hundreds of meters. Understanding associational effects could inform strategies for restoration or conservation; for example, planting of fir among existing natural regeneration could be concentrated in areas of low nutritional quality.