Balancing growth amidst salt stress - lifestyle perspectives from the extremophyte model Schrenkiella parvula.

Schrenkiella parvula, a leading extremophyte model in Brassicaceae, can grow and complete its lifecycle under multiple environmental stresses, including high salinity. Yet, the key physiological and structural traits underlying its stress-adapted lifestyle are unknown along with trade-offs when surviving salt stress at the expense of growth and reproduction. We aimed to identify the influential adaptive trait responses that lead to stress-resilient and uncompromised growth across developmental stages when treated with salt at levels known to inhibit growth in Arabidopsis and most crops. Its resilient growth was promoted by traits that synergistically allowed primary root growth in seedlings, the expansion of xylem vessels across the root-shoot continuum, and a high capacity to maintain tissue water levels by developing thicker succulent leaves while enabling photosynthesis during salt stress. A successful transition from vegetative to reproductive phase was initiated by salt-induced early flowering, resulting in viable seeds. Self-fertilization in salt-induced early flowering was dependent upon filament elongation in flowers otherwise aborted in the absence of salt during comparable plant ages. The maintenance of leaf water status promoting growth, and early flowering to ensure reproductive success in a changing environment, were among the most influential traits that contributed to the extremophytic lifestyle of S. parvula.

Life at the conservative end of the leaf economics spectrum: intergeneric variation in the allocation of phosphorus to biochemical fractions in species of Banksia (Proteaceae) and Hakea (Proteaceae).

In severely phosphorus (P)-impoverished environments, plants have evolved to use P very efficiently. Yet, it is unclear how P allocation in leaves contributes to their photosynthetic P-use efficiency (PPUE) and position along the leaf economics spectrum (LES). We address this question in 10 species of Banksia and Hakea, two highly P-efficient Proteaceae genera. We characterised traits in leaves of Banksia and Hakea associated with the LES: leaf mass per area, light-saturated photosynthetic rates, P and nitrogen concentrations, and PPUE. We also determined leaf P partitioning to five biochemical fractions (lipid, nucleic acid, metabolite, inorganic and residual P) and their possible association with the LES. For both genera, PPUE was negatively correlated with fractional allocation of P to lipids, but positively correlated with that to metabolites. For Banksia only, PPUE was negatively correlated with residual P, highlighting a strategy contrasting to that of Hakea. Phosphorus-allocation patterns significantly explained PPUE but were not linked to the resource acquisition vs resource conservation gradient defined by the LES. We conclude that distinct P-allocation patterns enable species from different genera to achieve high PPUE and discuss the implications of different P investments. We surmise that different LES axes representing different ecological strategies coexist in extremely P-impoverished environments.

Facilitative and competitive interactions between mycorrhizal and nonmycorrhizal plants in an extremely phosphorus-impoverished environment: role of ectomycorrhizal fungi and native oomycete pathogens in shaping species coexistence.

Nonmycorrhizal cluster root-forming species enhance the phosphorus (P) acquisition of mycorrhizal neighbours in P-impoverished megadiverse systems. However, whether mycorrhizal plants facilitate the defence of nonmycorrhizal plants against soil-borne pathogens, in return and via their symbiosis, remains unknown. We characterised growth and defence-related compounds in Banksia menziesii (nonmycorrhizal) and Eucalyptus todtiana (ectomycorrhizal, ECM) seedlings grown either in monoculture or mixture in a multifactorial glasshouse experiment involving ECM fungi and native oomycete pathogens. Roots of B. menziesii had higher levels of phytohormones (salicylic and jasmonic acids, jasmonoyl-isoleucine and 12-oxo-phytodienoic acid) than E. todtiana which further activated a salicylic acid-mediated defence response in roots of B. menziesii, but only in the presence of ECM fungi. We also found that B. menziesii induced a shift in the defence strategy of E. todtiana, from defence-related secondary metabolites (phenolic and flavonoid) towards induced phytohormone response pathways. We conclude that ECM fungi play a vital role in the interactions between mycorrhizal and nonmycorrhizal plants in a severely P-impoverished environment, by introducing a competitive component within the facilitation interaction between the two plant species with contrasting nutrient-acquisition strategies. This study sheds light on the interplay between beneficial and detrimental soil microbes that shape plant-plant interaction in severely nutrient-impoverished ecosystems.

Growing in phosphorus-impoverished habitats in south-western Australia: How general are phosphorus-acquisition and -allocation strategies among Proteaceae, Fabaceae and Myrtaceae species?

Numerous phosphorus (P)-acquisition and -utilisation strategies have evolved in plants growing in severely P-impoverished environments. Although these strategies have been well characterised for certain taxa, like Proteaceae, P-poor habitats are characterised by a high biodiversity, and we know little about how species in other families cope with P scarcity. We compared the P-acquisition and leaf P-allocation strategies of Fabaceae and Myrtaceae with those of Proteaceae growing in the same severely P-impoverished habitat. Myrtaceae and Fabaceae exhibited multiple P-acquisition strategies: P-mining by carboxylates or phosphatases, P uptake facilitated by carboxylate-releasing neighbours, and dependence on the elevated soil P availability after fire. Surprisingly, not all species showed high photosynthetic P-use efficiency (PPUE). Highly P-efficient species showed positive correlations between PPUE and the proportion of metabolite P (enzyme substrates), and negative correlations between PPUE and phospholipids (cellular membranes) and nucleic acid P (mostly ribosomal RNA), while we found no correlations in less P-efficient species. Overall, we found that Myrtaceae and Fabaceae used a wider range of strategies than Proteaceae to cope with P scarcity, at both the rhizosphere and leaf level. This knowledge is pivotal to better understand the mechanisms underlying plant survival in severely nutrient-impoverished biodiverse ecosystems.

Identification and expression analysis of Phosphate Transporter 1 (PHT1) genes in the highly phosphorus-use-efficient Hakea prostrata (Proteaceae).

Heavy and costly use of phosphorus (P) fertiliser is often needed to achieve high crop yields, but only a small amount of applied P fertiliser is available to most crop plants. Hakea prostrata (Proteaceae) is endemic to the P-impoverished landscape of southwest Australia and has several P-saving traits. We identified 16 members of the Phosphate Transporter 1 (PHT1) gene family (HpPHT1;1-HpPHT1;12d) in a long-read genome assembly of H. prostrata. Based on phylogenetics, sequence structure and expression patterns, we classified HpPHT1;1 as potentially involved in Pi uptake from soil and HpPHT1;8 and HpPHT1;9 as potentially involved in Pi uptake and root-to-shoot translocation. Three genes, HpPHT1;4, HpPHT1;6 and HpPHT1;8, lacked regulatory PHR1-binding sites (P1BS) in the promoter regions. Available expression data for HpPHT1;6 and HpPHT1;8 indicated they are not responsive to changes in P supply, potentially contributing to the high P sensitivity of H. prostrata. We also discovered a Proteaceae-specific clade of closely-spaced PHT1 genes that lacked conserved genetic architecture among genera, indicating an evolutionary hot spot within the genome. Overall, the genome assembly of H. prostrata provides a much-needed foundation for understanding the genetic mechanisms of novel adaptations to low P soils in southwest Australian plants.

Adenanthos species (Proteaceae) in phosphorus-impoverished environments use a variety of phosphorus-acquisition strategies and achieve high-phosphorus-use efficiency.

BACKGROUND AND AIMS: Soils in south-western Australia are severely phosphorus (P) impoverished, and plants in this region have evolved a variety of P-acquisition strategies. Phosphorus acquisition by Adenanthos cygnorum (Proteaceae) is facilitated by P-mobilizing neighbours which allows it to extend its range of habitats. However, we do not know if other Adenanthos species also exhibit a strategy based on facilitation for P acquisition in P-impoverished environments. METHODS: We collected leaf and soil samples of Adenanthosbarbiger, A. cuneatus, A.meisneri,A. obovatus, A. sericeus and Adenanthos sp. Whicher Range (G.J. Keighery 9736) growing in their natural habitats at different locations within the severely P-limited megadiverse environment of south-western Australia. Hydroponic experiments were conducted to collect the carboxylates exuded by cluster roots. Pot experiments in soil were carried out to measure rhizosheath phosphatase activity. KEY RESULTS: We found no evidence for facilitation of P uptake in any of the studied Adenanthos species. Like most Proteaceae, A. cuneatus, A. meisneri, A. obovatus, A. sericeus and Adenanthos sp. Whicher Range (G.J. Keighery 9736) expressed P-mining strategies, including the formation of cluster roots. Cluster roots of A. obovatus were less effective than those of the other four Adenanthos species. In contrast to what is known for most Proteaceae, we found no cluster roots for A. barbiger. This species probably expressed a post-fire P-acquisition strategy. All Adenanthos species used P highly efficiently for photosynthesis, like other Proteaceae in similar natural habitats. CONCLUSIONS: Adenanthos is the first genus of Proteaceae found to express multiple P-acquisition strategies. The diversity of P-acquisition strategies in these Proteaceae, coupled with similarly diverse strategies in Fabaceae and Myrtaceae, demonstrates that caution is needed in making family- or genus-wide extrapolations about the strategies exhibited in severely P-impoverished megadiverse ecosystems.

Phosphorus fractions in leaves.

Leaf phosphorus (P) comprises four major fractions: inorganic phosphate (Pi ), nucleic acids, phospholipids, P-containing metabolites and a residual fraction. In this review paper, we investigated whether allocation of P fractions varies among groups of terrestrial vascular plants, and is indicative of a species' strategy to use P efficiently. We found that as leaf total P concentration increases, the Pi fraction increases the most, without a plateau, while other fractions plateau. Variability of the concentrations of leaf P fractions is greatest among families > species(family) > regions > plant life forms. The percentage of total P allocated to nucleic acid-P (20-35%) and lipid-P (14-34%) varies less among families/species. High photosynthetic P-use efficiency is associated with low concentrations of all P fractions, and preferential allocation of P to metabolite-P and mesophyll cells. Sequential resorption of P from senescing leaves starts with Pi , followed by metabolite-P, and then other organic P fractions. Allocation of P to leaf P fractions varies with season. Leaf phytate concentrations vary considerably among species, associated with variation in photosynthesis and defence. Plasticity of P allocation to its fractions is important for acclimation to low soil P availability, and species-specific P allocation is needed for co-occurrence with other species.

Approaches to increase the validity of gene family identification using manual homology search tools.

Identifying homologs is an important process in the analysis of genetic patterns underlying traits and evolutionary relationships among species. Analysis of gene families is often used to form and support hypotheses on genetic patterns such as gene presence, absence, or functional divergence which underlie traits examined in functional studies. These analyses often require precise identification of all members in a targeted gene family. Manual pipelines where homology search and orthology assignment tools are used separately are the most common approach for identifying small gene families where accurate identification of all members is important. The ability to curate sequences between steps in manual pipelines allows for simple and precise identification of all possible gene family members. However, the validity of such manual pipeline analyses is often decreased by inappropriate approaches to homology searches including too relaxed or stringent statistical thresholds, inappropriate query sequences, homology classification based on sequence similarity alone, and low-quality proteome or genome sequences. In this article, we propose several approaches to mitigate these issues and allow for precise identification of gene family members and support for hypotheses linking genetic patterns to functional traits.

First report of Phytopythium vexans causing root rot on Macadamia integrifolia in China.

Soil-borne plant-pathogenic Phytopythium spp. can cause root rot and damping off on important plant species, resulting in serious economic loss. A survey in October 2021 identified soil-borne diseases occurring on Macadamia integrifolia in Yunnan Province, China. Microbes were isolated from necrotic roots of 23 trees with root rot symptoms by growing on cornmeal-based oomycete-selective 3P (Haas 1964) and P5APR (Jeffers and Martin, 1986) media at 24ºC in the dark for 7 days. Of the 56 single-hyphal isolates obtained, 18 were morphologically similar to Phytopythium vexans (van der Plaats-Niterink 1981; de Cock et al. 2015). Isolates LC04 and LC051 were selected for molecular analyses. The internal transcribed spacer (ITS) region and the cytochrome c oxidase subunit II (CoxII) gene were PCR-amplified using universal primers ITS1/ITS4 (White et al. 1990) and oomycete-specific primers Cox2-F/Cox2-RC4 (Choi et al. 2015), respectively. The PCR products were sequenced with the amplification primers and sequences were lodged in Genbank (Accession no. OM346742, OM415989 for ITS, OM453644, OM453643 for CoxII for isolates LC04 and LC051, respectively). The top BLAST hit in the Genbank nr database for all four sequences was Phytopythium vexans (>99% identity). A maximum-likelihood phylogenetic tree was constructed with analogous concatenated ITS and CoxII sequences from either type or voucher specimens of 13 Phytopythium species in the same phylogenetic clade as P. vexans (Table 1; Bala et. al 2010). Isolates LC04 and LC051 grouped most closely to P. vexans, with LC051 basal and sister to LC04 and P. vexans voucher specimen CBS119.80 with 100% support (Fig. 1). Millet seed inoculated with agar pieces colonized by P. vexans LC04 and LC51 was used to fulfill Koch's postulates (Li et al. 2015) in a completely randomized experimental design. Four 6-month-old M. integrifolia var. Keaau (660) seedlings were transplanted into pasteurized commercial potting mix containing 0.5% (w/w) inoculum. Plants were grown in free draining pots and watered once a day. At 14 days post-inoculation, roots were discolored compared to control plants inoculated with millet seed mixed with agar plugs lacking P. vexans (Fig. 2). By 30 days post-inoculation, infected roots were discolored with obvious decay and reduction in root system size. Control plants were symptomless. P. vexans was successfully re-isolated from two lesioned roots from each plant. The infection experiment was done twice, demonstrating that P. vexans LC04 and LC51 caused root disease on M. integrifolia. P. vexans causes root rot, damping-off, crown rot, stem rot or patch canker on economically important trees in many parts of the world, including seven plant species in China (Farr and Rossman 2022). This is the first report of pathogenic P. vexans on M. integrifolia in China. Reports of pathogenic P. vexans on multiple hosts in several parts of the world suggest it should be considered a quarantine risk and included in risk mitigation or pest management plans that include other species of Phytopythium, or species of Pythium or Phytophthora, to which P. vexans has many similarities (de Cock et al. 2015).

First report of Neofusicoccum parvum causing leaf spot disease on Macadamia integrifolia in China.

The macadamia industry is developing rapidly in China. A brown leaf spot disease was noted in six Macadamia integrifolia plantations in Lincang, Yunnan, in October 2021. Over 60% of trees sampled had brown leaf spot symptoms, among approx.15,000 trees planted in these areas. Lesions (3 to 5 mm dia.) were small round brown spots with yellow edges. Lesions on severely infected leaves were darker and larger, with irregular shape (8 to 10 mm long, 3 to 6 mm wide). About 10% of diseased leaves had lesions characterized by a shot hole surrounded by a yellow halo. Potential pathogens were isolated from four randomly-selected symptomatic leaves from each of the six plantations by cutting lesion edges into small pieces. The pieces were surface sterilized, placed onto water agar containing 100 ppm aureomycin and incubated for 5 days at 24°C in the dark. Subculturing microbial growth on potato dextrose agar produced single-hyphal isolates with white fluffy aerial mycelia that turned pale olivaceous gray after 4 to 5 days. In four randomly-selected cultures, conidia were single celled, hyaline, spindle shaped to oval, and measured 10.9 to 16.3 µm long and 4.0 to 6.2 µm wide (n = 50). These characteristics matched those of Neofusicoccum parvum (Pavlic et al. 2009). Isolate LC013 was randomly selected as a representative individual for molecular identification. Internal transcribed spacer (ITS; ITS1/ITS4 primers; White et al. 1990), beta-tubulin gene (tub2; BT2A/BT2B primers; Glass and Donaldson 1995) and translation elongation factor 1-alpha gene (tef1-α; EF1-728F/EF2 primers; Carbone and Kohn 1999; O'Donnell et al. 1998) regions were PCR amplified from genomic DNA. Sequences of the products were used to BLAST probe the type specimen nucleotide sequences in GenBank. The LC013 sequences (GenBank accessions OM392021 (ITS); OM453641 (tub2); OM567656 (tef1-α)) had >99% sequence identity with analogous sequences from the type specimen of N. parvum CBS 138823 (accessions AY236943 (ITS); AY236917 (tub2); AY236888 (tef1-α)). Isolate LC013 was sister to N. parvum type strain in a maximum-likelihood (ML) tree constructed from analogous concatenated ITS, tef1-α, and tub2 sequences of 27 species that are phylogenetically closely-related to LC013 based on the ITS single locus ML tree. Koch's postulates were tested twice with two isolates by wounding leaves of four 14-month-old M. integrifolia seedlings with a sterile needle and placing a 5-mm-diameter agar plug containing N. parvum on the wound site. PDA plugs alone were used as uninoculated controls. Leaves were covered with sealed bags to maintain >90% humidity for 24 hours. All plants were kept in the same glasshouse under natural conditions. Leaves of inoculated plants began to discolor at 5 days post-inoculation (dpi). Brown spot symptoms were observed at 9 dpi. Control plants were symptomless. N. parvum was re-isolated from leaf lesions of the infected plants, but not from control plants, thus fulfilling Koch's postulates. N. parvum is an aggressive pathogen that causes severe disease on important tree and woody species, including M. integrifolia (Liddle et al. 2019). In China, it has been reported to cause leaf spot disease on 26 plant species (Farr and Rossman 2022), but this is the first report of N. parvum causing leaf spot disease on M. integrifolia. Further investigation is required to estimate the importance of this pathogen to the macadamia industry in China.

Pangenomes as a Resource to Accelerate Breeding of Under-Utilised Crop Species.

Pangenomes are a rich resource to examine the genomic variation observed within a species or genera, supporting population genetics studies, with applications for the improvement of crop traits. Major crop species such as maize (Zea mays), rice (Oryza sativa), Brassica (Brassica spp.), and soybean (Glycine max) have had pangenomes constructed and released, and this has led to the discovery of valuable genes associated with disease resistance and yield components. However, pangenome data are not available for many less prominent crop species that are currently under-utilised. Despite many under-utilised species being important food sources in regional populations, the scarcity of genomic data for these species hinders their improvement. Here, we assess several under-utilised crops and review the pangenome approaches that could be used to build resources for their improvement. Many of these under-utilised crops are cultivated in arid or semi-arid environments, suggesting that novel genes related to drought tolerance may be identified and used for introgression into related major crop species. In addition, we discuss how previously collected data could be used to enrich pangenome functional analysis in genome-wide association studies (GWAS) based on studies in major crops. Considering the technological advances in genome sequencing, pangenome references for under-utilised species are becoming more obtainable, offering the opportunity to identify novel genes related to agro-morphological traits in these species.

Metabolic acclimation supports higher aluminium-induced secretion of citrate and malate in an aluminium-tolerant hybrid clone of Eucalyptus.

BACKGROUND: Eucalyptus is the main plantation wood species, mostly grown in aluminized acid soils. To understand the response of Eucalyptus clones to aluminum (Al) toxicity, the Al-tolerant Eucalyptus grandis × E. urophylla clone GL-9 (designated "G9") and the Al-sensitive E. urophylla clone GL-4 (designated "W4") were employed to investigate the production and secretion of citrate and malate by roots. RESULTS: Eucalyptus seedlings in hydroponics were exposed to the presence or absence of 4.4 mM Al at pH 4.0 for 24 h. The protein synthesis inhibitor cycloheximide (CHM) and anion channel blocker phenylglyoxal (PG) were applied to explore possible pathways involved in organic acid secretion. The secretion of malate and citrate was earlier and greater in G9 than in W4, corresponding to less Al accumulation in G9. The concentration of Al in G9 roots peaked after 1 h and decreased afterwards, corresponding with a rapid induction of malate secretion. A time-lag of about 6 h in citrate efflux in G9 was followed by robust secretion to support continuous Al-detoxification. Malate secretion alone may alleviate Al toxicity because the peaks of Al accumulation and malate secretion were simultaneous in W4, which did not secrete appreciable citrate. Enhanced activities of citrate synthase (CS) and phosphoenolpyruvate carboxylase (PEPC), and reduced activities of isocitrate dehydrogenase (IDH), aconitase (ACO) and malic enzyme (ME) were closely associated with the greater secretion of citrate in G9. PG effectively inhibited citrate and malate secretion in both Eucalyptus clones. CHM also inhibited malate and citrate secretion in G9, and citrate secretion in W4, but notably did not affect malate secretion in W4. CONCLUSIONS: G9 immediately secrete malate from roots, which had an initial effect on Al-detoxification, followed by time-delayed citrate secretion. Pre-existing anion channel protein first contributed to malate secretion, while synthesis of carrier protein appeared to be needed for citrate excretion. The changes of organic acid concentrations in response to Al can be achieved by enhanced CS and PEPC activities, but was supported by changes in the activities of other enzymes involved in organic acid metabolism. The above information may help to further explore genes related to Al-tolerance in Eucalyptus.

Foliar nutrient allocation patterns in Banksia attenuata and Banksia sessilis differing in growth rate and adaptation to low-phosphorus habitats.

BACKGROUND AND AIMS: Phosphorus (P) and nitrogen (N) are essential nutrients that frequently limit primary productivity in terrestrial ecosystems. Efficient use of these nutrients is important for plants growing in nutrient-poor environments. Plants generally reduce foliar P concentration in response to low soil P availability. We aimed to assess ecophysiological mechanisms and adaptive strategies for efficient use of P in Banksia attenuata (Proteaceae), naturally occurring on deep sand, and B. sessilis, occurring on shallow sand over laterite or limestone, by comparing the allocation of P among foliar P fractions. METHODS: We carried out pot experiments with slow-growing B. attenuata, which resprouts after fire, and faster growing opportunistic B. sessilis, which is killed by fire, on substrates with different P availability using a randomized complete block design. We measured leaf P and N concentrations, photosynthesis, leaf mass per area, relative growth rate and P allocated to major biochemical fractions in B. attenuata and B. sessilis. KEY RESULTS: The two species had similarly low foliar total P concentrations, but distinct patterns of P allocation to P-containing fractions. The foliar total N concentration of B. sessilis was greater than that of B. attenuata on all substrates. The foliar total P and N concentrations in both species decreased with decreasing P availability. The relative growth rate of both species was positively correlated with concentrations of both foliar nucleic acid P and total N, but there was no correlation with other P fractions. Faster growing B. sessilis allocated more P to nucleic acids than B. attenuata did, but other fractions were similar. CONCLUSIONS: The nutrient allocation patterns in faster growing opportunistic B. sessilis and slower growing B. attenuata revealed different strategies in response to soil P availability which matched their contrasting growth strategy.

Direct to Consumer Telemedicine: Is Healthcare From Home Best?

Direct-to-consumer (DTC) telemedicine is increasingly popular and enables patients to obtain medical advice and treatment via electronic media (e.g., computer, telephone, or smartphone) without a prior doctor-patient relationship. Convenience, accessibility, and home delivery make DTC telemedicine attractive to patients. Concerns about DTC telemedicine include: a lack of regulation, transparency, and an established patient-provider relationship (physician and pharmacist). In future, researchers, providers, and insurers need to better understand the concerns and challenges that this new form of healthcare poses.

Responses of foliar phosphorus fractions to soil age are diverse along a 2 Myr dune chronosequence.

Plants respond to soil phosphorus (P) availability by adjusting leaf P among inorganic P (Pi) and organic P fractions (nucleic acids, phospholipids, small metabolites and a residual fraction). We tested whether phylogenetically divergent plants in a biodiversity hotspot similarly adjust leaf P allocation in response to P limitation by sampling along a 2 Myr chronosequence in southwestern Australia where nitrogen (N) limitation transitions to P limitation with increasing soil age. Total P and N, and P allocated to five chemical fractions were determined for photosynthetic organs from Melaleuca systena (Myrtaceae), Acacia rostellifera (Fabaceae) and Hakea prostrata (Proteaceae). Soil characteristics were also determined. Acacia rostellifera maintained phyllode total P and N concentrations at c. 0.5 and 16 mg g-1 DW, respectively, with a constant P-allocation pattern along the chronosequence. H. prostrata leaves allocated less P to Pi, phospholipids and nucleic acids with increasing soil age, while leaf N concentration was constant. M. systena had the greatest variation in allocating leaf P, whereas leaf N concentration decreased 20% along the chronosequence. Variation in P-allocation patterns was only partially conserved among species along the chronosequence. Such variation could have an impact on species distribution and contribute to species richness in P-limited environments.

Ceragenins are active against drug-resistant Candida auris clinical isolates in planktonic and biofilm forms.

Background: Candida auris has emerged as a serious threat to human health. Of particular concern are the resistance profiles of many clinical isolates, with some being resistant to multiple classes of antifungals. Objectives: Measure susceptibilities of C. auris isolates, in planktonic and biofilm forms, to ceragenins (CSAs). Determine the effectiveness of selected ceragenins in gel and cream formulations in eradicating fungal infections in tissue explants. Materials and methods: A collection of 100 C. auris isolates available at CDC was screened for susceptibility to a lead ceragenin. A smaller collection was used to characterize antifungal activities of other ceragenins against organisms in planktonic and biofilm forms. Effects of ceragenins on fungal cells and biofilms were observed via microscopy. An ex vivo model of mucosal fungal infection was used to evaluate formulated forms of lead ceragenins. Results: Lead ceragenins displayed activities comparable to those of known antifungal agents against C. auris isolates with MICs of 0.5-8 mg/L and minimum fungicidal concentrations (MFCs) of 2-64 mg/L. No cross-resistance with other antifungals was observed. Fungal cell morphology was altered in response to ceragenin treatment. Ceragenins exhibited activity against sessile organisms in biofilms. Gel and cream formulations including 2% CSA-44 or CSA-131 resulted in reductions of over 4 logs against established fungal infections in ex vivo mucosal tissues. Conclusions: Ceragenins demonstrated activity against C. auris, suggesting that these compounds warrant further study to determine whether they can be used for topical applications to skin and mucosal tissues for treatment of infections with C. auris and other fungi.

Heartwood-specific transcriptome and metabolite signatures of tropical sandalwood (Santalum album) reveal the final step of (Z)-santalol fragrance biosynthesis.

Tropical sandalwood (Santalum album) produces one of the world's most highly prized fragrances, which is extracted from mature heartwood. However, in some places such as southern India, natural populations of this slow-growing tree are threatened by over-exploitation. Sandalwood oil contains four major and fragrance-defining sesquiterpenols: (Z)-α-santalol, (Z)-ß-santalol, (Z)-epi-ß-santalol and (Z)-α-exo-bergamotol. The first committed step in their biosynthesis is catalyzed by a multi-product santalene/bergamotene synthase. Sandalwood cytochromes P450 of the CYP76F sub-family were recently shown to hydroxylate santalenes and bergamotene; however, these enzymes produced mostly (E)-santalols and (E)-α-exo-bergamotol. We hypothesized that different santalene/bergamotene hydroxylases evolved in S. album to stereo-selectively produce (E)- or (Z)-sesquiterpenols, and that genes encoding (Z)-specific P450s contribute to sandalwood oil formation if co-expressed in the heartwood with upstream genes of sesquiterpene biosynthesis. This hypothesis was validated by the discovery of a heartwood-specific transcriptome signature for sesquiterpenoid biosynthesis, including highly expressed SaCYP736A167 transcripts. We characterized SaCYP736A167 as a multi-substrate P450, which stereo-selectively produces (Z)-α-santalol, (Z)-ß-santalol, (Z)-epi-ß-santalol and (Z)-α-exo-bergamotol, matching authentic sandalwood oil. This work completes the discovery of the biosynthetic enzymes of key components of sandalwood fragrance, and highlights the evolutionary diversification of stereo-selective P450s in sesquiterpenoid biosynthesis. Bioengineering of microbial systems using SaCYP736A167, combined with santalene/bergamotene synthase, has potential for development of alternative industrial production systems for sandalwood oil fragrances.

Tight control of sulfur assimilation: an adaptive mechanism for a plant from a severely phosphorus-impoverished habitat.

Hakea prostrata (Proteaceae) has evolved in extremely phosphorus (P)-impoverished habitats. Unlike species that evolved in P-richer environments, it tightly controls its nitrogen (N) acquisition, matching its low protein concentration, and thus limiting its P requirement for ribosomal RNA (rRNA). Protein is a major sink for sulfur (S), but the link between low protein concentrations and S metabolism in H. prostrata is unknown, although this is pivotal for understanding this species' supreme adaptation to P-impoverished soils. Plants were grown at different sulfate supplies for 5 wk and used for nutrient and metabolite analyses. Total S content in H. prostrata was unchanged with increasing S supply, in sharp contrast with species that typically evolved in environments where P is not a major limiting nutrient. Unlike H. prostrata, other plants typically store excess available sulfate in vacuoles. Like other species, S-starved H. prostrata accumulated arginine, lysine and O-acetylserine, indicating S deficiency. Hakea prostrata tightly controls its S acquisition to match its low protein concentration and low demand for rRNA, and thus P, the largest organic P pool in leaves. We conclude that the tight control of S acquisition, like that of N, helps H. prostrata to survive in P-impoverished environments.

Tight control of nitrate acquisition in a plant species that evolved in an extremely phosphorus-impoverished environment.

Hakea prostrata (Proteaceae) has evolved in an extremely phosphorus (P)-limited environment. This species exhibits an exceptionally low ribosomal RNA (rRNA) and low protein and nitrogen (N) concentration in its leaves. Little is known about the N requirement of this species and its link to P metabolism, despite this being the key to understanding how it functions with a minimal P budget. H. prostrata plants were grown with various N supplies. Metabolite and elemental analyses were performed to determine its N requirement. H. prostrata maintained its organ N content and concentration at a set point, independent of a 25-fold difference nitrate supplies. This is in sharp contrast to plants that are typically studied, which take up and store excess nitrate. Plants grown without nitrate had lower leaf chlorophyll and carotenoid concentrations, indicating N deficiency. However, H. prostrata plants at low or high nitrate availability had the same photosynthetic pigment levels and hence were not physiologically compromised by the treatments. The tight control of nitrate acquisition in H. prostrata retains protein at a very low level, which results in a low demand for rRNA and P. We surmise that the constrained nitrate acquisition is an adaptation to severely P-impoverished soils.

Sensitivity of jarrah (Eucalyptus marginata) to phosphate, phosphite, and arsenate pulses as influenced by fungal symbiotic associations.

Many plant species adapted to P-impoverished soils, including jarrah (Eucalyptus marginata), develop toxicity symptoms when exposed to high doses of phosphate (Pi) and its analogs such as phosphite (Phi) and arsenate (AsV). The present study was undertaken to investigate the effects of fungal symbionts Scutellospora calospora, Scleroderma sp., and Austroboletus occidentalis on the response of jarrah to highly toxic pulses (1.5 mmol kg(-1) soil) of Pi, Phi, and AsV. S. calospora formed an arbuscular mycorrhizal (AM) symbiosis while both Scleroderma sp. and A. occidentalis established a non-colonizing symbiosis with jarrah plants. All these interactions significantly improved jarrah growth and Pi uptake under P-limiting conditions. The AM fungal colonization naturally declines in AM-eucalypt symbioses after 2-3 months; however, in the present study, the high Pi pulse inhibited the decline of AM fungal colonization in jarrah. Four weeks after exposure to the Pi pulse, plants inoculated with S. calospora had significantly lower toxicity symptoms compared to non-mycorrhizal (NM) plants, and all fungal treatments induced tolerance against Phi toxicity in jarrah. However, no tolerance was observed for AsV-treated plants even though all inoculated plants had significantly lower shoot As concentrations than the NM plants. The transcript profile of five jarrah high-affinity phosphate transporter (PHT1 family) genes in roots was not altered in response to any of the fungal species tested. Interestingly, plants exposed to high Pi supplies for 1 day did not have reduced transcript levels for any of the five PHT1 genes in roots, and transcript abundance of four PHT1 genes actually increased. It is therefore suggested that jarrah, and perhaps other P-sensitive perennial species, respond positively to Pi available in the soil solution through increasing rather than decreasing the expression of selected PHT1 genes. Furthermore, Scleroderma sp. can be considered as a fungus with dual functional capacity capable of forming both ectomycorrhizal and non-colonizing associations, where both pathways are always accompanied by evident growth and nutritional benefits.