BACKGROUND:Many parasitic plants of the genera Striga and Cuscuta inflict huge agricultural damage worldwide. To form and maintain a connection with a host plant, parasitic plants deploy virulence factors (VFs) that interact with host biology. They possess a secretome that represents the complement of proteins secreted from cells and like other plant parasites such as fungi, bacteria or nematodes, some secreted proteins represent VFs crucial to successful host colonisation. Understanding the genome-wide complement of putative secreted proteins from parasitic plants, and their expression during host invasion, will advance understanding of virulence mechanisms used by parasitic plants to suppress/evade host immune responses and to establish and maintain a parasite-host interaction.RESULTS:We conducted a comparative analysis of the secretomes of root (Striga spp.) and shoot (Cuscuta spp.) parasitic plants, to enable prediction of candidate VFs. Using orthogroup clustering and protein domain analyses we identified gene families/functional annotations common to both Striga and Cuscuta species that were not present in their closest non-parasitic relatives (e.g. strictosidine synthase like enzymes), or specific to either the Striga or Cuscuta secretomes. For example, Striga secretomes were strongly associated with 'PAR1' protein domains. These were rare in the Cuscuta secretomes but an abundance of 'GMC oxidoreductase' domains were found, that were not present in the Striga secretomes. We then conducted transcriptional profiling of genes encoding putatively secreted proteins for the most agriculturally damaging root parasitic weed of cereals, S. hermonthica. A significant portion of the Striga-specific secretome set was differentially expressed during parasitism, which we probed further to identify genes following a 'wave-like' expression pattern peaking in the early penetration stage of infection. We identified 39 genes encoding putative VFs with functions such as cell wall modification, immune suppression, protease, kinase, or peroxidase activities, that are excellent candidates for future functional studies.CONCLUSIONS:Our study represents a comprehensive secretome analysis among parasitic plants and revealed both similarities and differences in candidate VFs between Striga and Cuscuta species. This knowledge is crucial for the development of new management strategies and delaying the evolution of virulence in parasitic weeds.

Summary Parasites have evolved proteins, Virulence Factors (VFs), that facilitate plant colonization, yet VFs mediating parasitic plant-host interactions are poorly understood. Striga hermonthica is an obligate, root-parasitic plant of cereal hosts in sub-Saharan Africa, causing devastating yield losses. Understanding the molecular nature and allelic variation of VFs in S. hermonthica is essential for breeding resistance and delaying the evolution of parasite virulence. We assembled the S. hermonthica genome and identified secreted proteins by in silico prediction. Pooled sequencing of parasites growing on a susceptible and a strongly resistant rice host allowed us to scan for loci where selection imposed by the resistant host had elevated the frequency of alleles contributing to successful colonisation. Thirty-eight putatively secreted VFs had extremely different allele frequencies with functions including host cell wall modification, protease inhibitors, oxidoreductase and kinase activities. These candidate loci had significantly higher Tajima’s D than the genomic background, consistent with balancing selection. Our results reveal diverse strategies used by S. hermonthica to overcome different layers of host resistance. Understanding the maintenance of variation at virulence loci by balancing selection will be critical to managing the evolution of virulence as a part of a sustainable control strategy.

SUMMARYDeciphering signalling pathways is essential to understanding how organisms respond to environmental cues but elucidating how these signalling pathways evolve in new environments is less clear.1,2 Most plants, for example, monitor multiple environmental cues to optimize the time and place to germinate. Some root parasitic plants, however, germinate in response to small molecules like strigolactones (SLs) emanating from host roots3,4 whilst a number of ephemeral weeds germinate in response to chemicals called karrikins (KARs) released after a forest fire.5,6 Although these species represent distinct clades, they use the same HYPOSENSITIVE TO LIGHT/KARRIKIN INSENSITIVE 2 (HTL/KAI2) signalling pathway to perceive strigolactones or karrikins, which suggests convergent evolution.3,5 Because specialist lifestyles are derived traits, it is not clear if HTL/KAI2 signalling in these species evolved from a specific germination-signalling pathway or whether this pathway had other functions that were co-opted for specialist germination circumstances. Here, we show HTL/KAI2 signalling in Arabidopsis bypasses the light requirement for germination. In part, this is because the HTL/KAI2 downstream component, SMAX1 impinges on PHYTOCHROME INTERACTING FACTOR 1/PHYTOCHROME INTERACTING FACTOR 3-LIKE 5 (PIF1/PIL5)-regulated hormone response pathways conducive to germination. We identified Arabidopsis accessions that can germinate in the dark, which had altered expression of HTL/KAI2 signalling components, suggesting that divergence in this signalling pathway occurs in nature. Moreover, Arabidopsis HTL/KAI2-regulated gene signatures were observed in germinating Striga seed. The ability of HTL/KAI2 signalling to substitute for light advances an explanation for how some specialist plants evolved their underground germination behaviour in response to specific environments.

Summary Parasites have evolved proteins, virulence factors (VFs), that facilitate plant colonisation, however VFs mediating parasitic plant–host interactions are poorly understood. Striga hermonthica is an obligate, root‐parasitic plant of cereal hosts in sub‐Saharan Africa, causing devastating yield losses. Understanding the molecular nature and allelic variation of VFs in S. hermonthica is essential for breeding resistance and delaying the evolution of parasite virulence. We assembled the S. hermonthica genome and identified secreted proteins using in silico prediction. Pooled sequencing of parasites growing on a susceptible and a strongly resistant rice host allowed us to scan for loci where selection imposed by the resistant host had elevated the frequency of alleles contributing to successful colonisation. Thirty‐eight putatively secreted VFs had very different allele frequencies with functions including host cell wall modification, protease or protease inhibitor and kinase activities. These candidate loci had significantly higher Tajima's D than the genomic background, consistent with balancing selection. Our results reveal diverse strategies used by S. hermonthica to overcome different layers of host resistance. Understanding the maintenance of variation at virulence loci by balancing selection will be critical to managing the evolution of virulence as part of a sustainable control strategy.

Deciphering signalling pathways is essential to understanding how organisms respond to environmental cues but elucidating how these signalling pathways evolve in new environments is less clear.[1][1],[2][2] Most plants, for example, monitor multiple environmental cues to optimize the time and place to germinate. Some root parasitic plants, however, germinate in response to small molecules like strigolactones (SLs) emanating from host roots[3][3],[4][4] whilst a number of ephemeral weeds germinate in response to chemicals called karrikins (KARs) released after a forest fire.[5][5],[6][6] Although these species represent distinct clades, they use the same HYPOSENSITIVE TO LIGHT/KARRIKIN INSENSITIVE 2 (HTL/KAI2) signalling pathway to perceive strigolactones or karrikins, which suggests convergent evolution.[3][3],[5][5] Because specialist lifestyles are derived traits, it is not clear if HTL/KAI2 signalling in these species evolved from a specific germination-signalling pathway or whether this pathway had other functions that were co-opted for specialist germination circumstances. Here, we show HTL/KAI2 signalling in Arabidopsis bypasses the light requirement for germination. In part, this is because the HTL/KAI2 downstream component, SMAX1 impinges on PHYTOCHROME INTERACTING FACTOR 1/PHYTOCHROME INTERACTING FACTOR 3-LIKE 5 (PIF1/PIL5)-regulated hormone response pathways conducive to germination. We identified Arabidopsis accessions that can germinate in the dark, which had altered expression of HTL/KAI2 signalling components, suggesting that divergence in this signalling pathway occurs in nature. Moreover, Arabidopsis HTL/KAI2 -regulated gene signatures were observed in germinating Striga seed. The ability of HTL/KAI2 signalling to substitute for light advances an explanation for how some specialist plants evolved their underground germination behaviour in response to specific environments. ### Competing Interest Statement The authors have declared no competing interest. [1]: #ref-1 [2]: #ref-2 [3]: #ref-3 [4]: #ref-4 [5]: #ref-5 [6]: #ref-6

The parasitic weed genus Striga causes huge losses to crop production in sub-Saharan Africa, estimated to be in excess of $7 billion per year, affecting subsistence farmers who frequently lack access to novel technologies proposed for control. Effective Striga management therefore requires the development of strategies utilising existing cultural and management practices. We report a multi-year, landscape-scale monitoring project for Striga asiatica in the mid-west of Madagascar, undertaken over 2019-2020 with the aims of examining cultural, climatic and edaphic factors currently driving abundance and distribution. Long-distance transects were established across the middle-west region of Madagascar, over which Striga asiatica abundance in fields was estimated. Analysis of the data highlights the importance of crop variety and legumes in driving Striga density. Moreover, the dataset revealed significant effect of precipitation seasonality, mean temperature and altitude in determining abundance. A composite management index indicated the effect of a range of cultural practices on changes in Striga abundance. The findings support the assertion that single measures are not sufficient for the effective, long-term management of Striga. Furthermore, the composite score has potential as a significant guide of ISM control beyond the geographic range of this study.

Parasitic plants connect to the vasculature of a host plant and take part or all of the water, nutrients, and assimilates they need to complete their life cycle. Parasitic plants represent a unique model for the evolution of intra-kingdom parasitism with intriguing research questions such as how plants were able to evolve the ability to parasitize other plants. This parasitic lifestyle required the evolution of host detection, host attachment, host exploitation, and host defense suppression strategies. The elucidation of these strategies and the underlying mechanisms has been greatly facilitated by the advent of model species, molecular and genomic tools, and omics approaches such as transcriptomics and comparative genomics and this has launched parasitic plant research into the 21st century. The scientific data generated are helping us to gradually unravel how parasitism in plants has evolved. However, parasitic plants are not just an interesting basic research model. Due to their parasitic lifestyle, many parasitic plant species represent important agricultural weeds, such as the witchweeds, broomrapes, dodders, and mistletoes. Fundamental knowledge of the mechanisms underlying parasitism will also contribute to new approaches to create resistance in crops and develop new control measures for this important agricultural problem. This Focus Issue on Parasitic Plants addresses some of the most important advances and new landmarks in the field. In addition to commissioned updates by experts in their respective topics, several research articles highlight recent accomplishments in these areas. The name “parasitic plant” illustrates that these plants grow at the expense of their hosts and can cause problems in agriculture, as will be further discussed below. But parasitic plants are also an essential component of biodiverse ecosystems. In two updates, Casadesús and Munné-Bosch (2021) and T e sitel et al. (2021) describe the role of parasitic plants in natural ecosystems. Casadesús and MunnéBosch (2021) highlight the role of holoparasitic plant–host interactions (such as those between Cytinus hypocistis and various shrubs of the genus Cistus) in shaping natural Mediterranean ecosystems. T e sitel et al. (2021) discuss how parasitic plants can also positively influence the growth and/ or reproductive output of their host or other, nonhost, organisms and that they can have positive effects on community structure and ecosystem processes. T e sitel et al. (2021) also review the roles that parasitic plants may have as food or medicinal resources or for aesthetic and cultural purposes. During the early days of genomics, it was assumed that parasitic plants must have acquired pathogenicity genes through horizontal gene transfer. Surprisingly, in the last decade or so it also became clear that pathogenicity evolved from extant physiological processes also present in nonparasitic plant species. This Focus Issue contains several examples of the recruitment by parasitic plants of genes involved in these extant physiological processes, to facilitate aspects of the parasitic process. A prime example is the coordination of the parasitic plant lifecycle with that of the host, aided by a number of host detection/host response mechanisms that have evolved in parasitic plant species. In the update by Brun et al. (2021), the molecular mechanisms underlying two of these—host-dependent seed germination and haustorium formation—are reviewed. The key process Ed it o ri al

AbstractInfestations by the parasitic weed genus Striga result in significant losses to cereal crop yields across sub‐Saharan Africa. The problem disproportionately affects subsistence farmers who frequently lack access to novel technologies. Effective Striga management therefore requires the development of strategies utilizing existing cultural management practices. We report a multiyear, landscape‐scale monitoring project for Striga asiatica in the mid‐west of Madagascar, undertaken over 2019–2020 with the aims of examining cultural, climatic, and edaphic factors currently driving abundance and distribution. Long‐distance transects were established across the middle‐west region of Madagascar, over which S. asiatica abundance in fields was estimated. Analysis of the data highlights the importance of crop variety and legumes in driving Striga density. Moreover, the dataset revealed significant effect of precipitation seasonality, mean temperature, and altitude in determining abundance. A composite management index indicated the effect of a range of cultural practices on changes in Striga abundance. The findings support the assertion that single measures are not sufficient for the effective, long‐term management of Striga. Furthermore, the composite score has potential as a significant guide of integrated Striga management beyond the geographic range of this study.

Parasitic plant infestations dramatically reduce the yield of many major food crops of sub-Saharan Africa and pose a serious threat to food security on that continent 1 . The first committed step of a successful infestation is the germination of parasite seeds primarily in response to a group of related small-molecule hormones called strigolactones (SLs), which are emitted by host roots 2 . Despite the important role of SLs, it is not clear how host-derived SLs germinate parasitic plants. In contrast, gibberellins (GA) acts as the dominant hormone for stimulation of germination in non-parasitic plant species by inhibiting a set of DELLA repressors 3 . Here, we show that expression of SL receptors from the parasitic plant Striga hermonthica in the presence of SLs circumvents the GA requirement for germination of Arabidopsis thaliana seed. Striga receptors co-opt and enhance signalling through the HYPOSENSITIVE TO LIGHT/KARRIKIN INSENSITIVE 2 (AtHTL/KAI2) pathway, which normally plays a rudimentary role in Arabidopsis seed germination 4 , 5 . AtHTL/KAI2 negatively controls the SUPPRESSOR OF MAX2 1 (SMAX1) protein 5 , and loss of SMAX1 function allows germination in the presence of DELLA repressors. Our data suggest that ligand-dependent inactivation of SMAX1 in Striga and Arabidopsis can bypass GA-dependent germination in these species.

The parasitic weed genusStrigacauses huge losses to crop production in sub-Saharan Africa, estimated to be in excess of $7 billion per year. There is a paucity of reliable distribution data forStriga; however, such data are urgently needed to understand current drivers, better target control efforts, as well as to predict future risks. To address this, we developed a methodology to enable rapid, large-scale monitoring ofStrigapopulations. We used this approach to uncover the factors that currently drive the abundance and distribution ofStriga asiaticain Madagascar. Two long-distance transects were established across the middle-west region of Madagascar in whichS. asiaticaabundance in fields adjacent to the road was estimated. Management, crop structure and soil data were also collected. Analysis of the data suggests that crop variety, companion crop and previous crop were correlated withStrigadensity. A positive relationship between within-fieldStrigadensity and the density of the nearest neighbouring fields indicates that spatial configuration and connectivity of suitable habitats is also important in determiningStrigaspread. Our results demonstrate that we are able to capture distribution and management data forStrigadensity at a landscape scale and use this to understand the ecological and agronomic drivers of abundance. The importance of crop varieties and cropping patterns is significant, as these are key socio-economic elements of Malagasy cropping practices. Therefore, they have the potential to be promoted as readily available control options, rather than novel technologies requiring introduction.

Striga is a genus of parasitic plants that poses great danger to livelihoods of millions of smallholder farmers in sub-Saharan Africa by limiting production of staple cereals. The parasite attaches to the roots of the crop and establishes a vascular connection with the host’s xylem vessels in order to access water, organic and inorganic nutrients, leading to stunted growth and death of the infected plant. Mechanisms underpinning host–parasite interactions are not clearly understood making well-informed strategies for control of the parasite difficult. To facilitate studies of Striga–host interactions for molecular and genetic studies, we: (i) established an efficient protocol for growing Striga from seeds in tissue culture; (ii) optimized protocols for its regeneration through direct organogenesis and somatic embryogenesis, and (iii) determined the effect of co-culturing host and parasite in the same culture media. We found the best auxin and cytokinin concentrations to be: 10.7 μM naphthaleneacetic acid (NAA) and 2.2 μM 6-benzylaminopurine (BAP) for embryogenic callus regeneration and 1.1–4.4 μM BAP without NAA for shoot multiplication. While seedling, stem and leaf explants induced callus with the same frequency, seed radicles did not produce any callus. Unexpectedly, we found that when Striga callus was added in rice growing on culture media, the parasite adversely affected the host through formation of lesions on leaves and resulted in less shoot induction from callus in the parasite. Techniques described in this study will enhance further understanding of Striga–host interactions. Highly efficient protocols for tissue culture of the obligate parasitic plant Striga hermonthica and detection of host phytotoxicity by the parasite on culture media.

In the version of this Perspective originally published, 'acidification' was incorrectly spelt as 'adification' in Fig. 4. This has now been corrected.

In the version of this Perspective originally published, 'acidification' was incorrectly spelt as 'adification' in Fig. 4. This has now been corrected. © 2018 Macmillan Publishers Limited, part of Springer Nature. All rights reserved.

Restricting future global temperature increase to 2°C or less requires the adoption of negative emissions technologies for carbon capture and storage. We review the potential for deployment of enhanced weathering (EW), via the application of crushed reactive silicate rocks (such as basalt), on over 680 million hectares of tropical agricultural and tree plantations to offset fossil fuel CO 2 emissions. Warm tropical climates and productive crops will substantially enhance weathering rates, with potential co-benefits including decreased soil acidification and increased phosphorus supply promoting higher crop yields sparing forest for conservation, and reduced cultural eutrophication. Potential pitfalls include the impacts of mining operations on deforestation, producing the energy to crush and transport silicates and the erosion of silicates into rivers and coral reefs that increases inorganic turbidity, sedimentation and pH, with unknown impacts for biodiversity. We identify nine priority research areas for untapping the potential of EW in the tropics, including effectiveness of tropical agriculture at EW for major crops in relation to particle sizes and soil types, impacts on human health, and effects on farmland, adjacent forest and stream-water biodiversity.

The parasitic weeds Striga asiatica and Striga hermonthica cause devastating yield losses to upland rice in Africa. Little is known about genetic variation in host resistance and tolerance across rice genotypes, in relation to virulence differences across Striga species and ecotypes.Diverse rice genotypes were phenotyped for the above traits in S. asiatica-(Tanzania) and S. hermonthica-infested fields (Kenya and Uganda) and under controlled conditions.New rice genotypes with either ecotype-specific or broad-spectrum resistance were identified. Resistance identified in the field was confirmed under controlled conditions, providing evidence that resistance was largely genetically determined. Striga-resistant genotypes contributed to yield security under Striga-infested conditions, although grain yield was also determined by the genotype-specific yield potential and tolerance. Tolerance, the physiological mechanism mitigating Striga effects on host growth and physiology, was unrelated to resistance, implying that any combination of high, medium or low levels of these traits can be found across rice genotypes.Striga virulence varies across species and ecotypes. The extent of Striga-induced host damage results from the interaction between parasite virulence and genetically determined levels of host-plant resistance and tolerance. These novel findings support the need for predictive breeding strategies based on knowledge of host resistance and parasite virulence.

Background: Connections between mature trees and seedlings via ectomycorrhizal (EcM) hyphal networks existing in dipterocarp-dominated tropical rain forests of South-east Asia could have strong implications for seedling growth and survival and the maintenance of high diversity in such forests. Aim: To test whether EcM hyphal network connections are important for the growth and survival of dipterocarp seedlings. Methods: We conducted four independent experiments that prevented contact of experimental seedlings with an EcM network by using a series of fine meshes and/or plastic barriers. We measured the growth and survival (and foliar δ13C in one experiment) of seedlings of six dipterocarp species over intervals ranging from 11 to 29 months. Results: Seedling growth (diameter, height or leaf number) was unaffected by exclusion from the EcM network in three experiments and there were no differences in foliar δ13C values in the fourth. Seedling survival was reduced following exclusion from the EcM network in one experiment. Our results give little support to the hypothesis that dipterocarp seedlings growing in the shaded forest understorey benefit from being connected, through a common EcM network, to surrounding trees. Conclusions: We suggest that our negative results, in contrast to studies conducted in low diversity boreo-temperate or tropical forests, are due to these high diversity forests lacking host species-specific EcM fungi and therefore providing little opportunity for adaptive support of seedlings via hyphal networks.

The parasitic weeds Striga asiatica and Striga hermonthica cause high yield losses in rain-fed upland rice in Africa. Two resistance classes (pre- and post-attachment) and several resistant genotypes have been identified among NERICA (New Rice for Africa) cultivars under laboratory conditions (in vitro) previously. However, little is known about expression of this resistance under field conditions. Here we investigated (1) whether resistance exhibited under controlled conditions would express under representative Striga-infested field conditions, and (2) whether NERICA cultivars would achieve relatively good grain yields under Striga-infested conditions. Twenty-five rice cultivars, including all 18 upland NERICA cultivars, were screened in S. asiatica-infested (in Tanzania) and S. hermonthica-infested (in Kenya) fields during two seasons. Additionally, a selection of cultivars was tested in vitro, in mini-rhizotron systems. For the first time, resistance observed under controlled conditions was confirmed in the field for NERICA-2, 5, -10 and -17 (against S. asiatica) and NERICA-1 to -5, -10, -12, -13 and -17 (against S. hermonthica). Despite high Striga-infestation levels, yields of around 1.8 t ha(-1) were obtained with NERICA-1, -9 and -10 (in the S. asiatica-infested field) and around 1.4t ha(-1) with NERICA-3, -4, -8, -12 and -13 (in the S. hermonthica-infested field). In addition, potential levels of tolerance were identified in vitro, in NERICA-1, -17 and -9 (S. asiatica) and in NERICA-1, -17 and -10 (S. hermonthica). These findings are highly relevant to rice agronomists and breeders and molecular geneticists working on Striga resistance. In addition, cultivars combining broad-spectrum resistance with good grain yields in Striga-infested fields can be recommended to rice farmers in Striga-prone areas. (C) 2014 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY license (http://creativecommons.org/licenses/by/3.0/).

Resistance-inducing chemicals can offer broad-spectrum disease protection in crops, but can also affect plant growth and interactions with plant-beneficial microbes. We have evaluated different application methods of β-aminobutyric acid (BABA) and jasmonic acid (JA) for long-lasting induced resistance in tomato against Botrytis cinerea. In addition, we have studied nontarget effects on plant growth and root colonization by arbuscular mycorrhizal fungi (AMF). Germinating seeds for 1 week in BABA- or JA-containing solutions promoted seed germination efficiency, did not affect plant growth, and induced resistance in 4-week-old plants. When formulating BABA and JA in carboxymethyl cellulose seed coating, only BABA was able to induce resistance in 4-week-old plants. Root treatment of 1-week-old seedlings with BABA or JA also induced resistance in 4-week-old plants. However, this seedling treatment repressed plant growth at higher concentrations of the chemicals, which was particularly pronounced in hydroponically grown plants after BABA treatment. Both seed coating with BABA, and seedling treatments with BABA or JA, did not affect AMF root colonization in soil-grown tomato. Our study has identified commercially feasible application methods of BABA and JA, which induce durable disease resistance in tomato without concurrent impacts on plant growth or colonization by plant-beneficial AMF.

The parasitic weeds Striga asiatica and Striga hermonthica cause high yield losses in rain-fed upland rice in Africa. Two resistance classes (preand post-attachment) and several resistant genotypes have been identified among NERICA (New Rice for Africa) cultivars under laboratory conditions (in vitro) previously. However, little is known about expression of this resistance under field conditions. Here we investigated (1) whether resistance exhibited under controlled conditions would express under representative Strigainfested field conditions, and (2) whether NERICA cultivars would achieve relatively good grain yields under Striga-infested conditions. Twenty-five rice cultivars, including all 18 upland NERICA cultivars, were screened in S. asiatica-infested (in Tanzania) and S. hermonthica-infested (in Kenya) fields during two seasons. Additionally, a selection of cultivars was tested in vitro, in mini-rhizotron systems. For the first time, resistance observed under controlled conditions was confirmed in the field for NERICA-2, 5, -10 and -17 (against S. asiatica) and NERICA-1 to -5, -10, -12, -13 and -17 (against S. hermonthica). Despite high Striga-infestation levels, yields of around 1.8 t ha−1 were obtained with NERICA-1, -9 and -10 (in the S. asiatica-infested field) and around 1.4 t ha−1 with NERICA-3, -4, -8, -12 and -13 (in the S. hermonthica-infested field). In addition, potential levels of tolerance were identified in vitro, in NERICA-1, -17 and -9 (S. asiatica) and in NERICA-1, -17 and -10 (S. hermonthica). These findings are highly relevant to rice agronomists and breeders and molecular geneticists working on Striga resistance. In addition, cultivars combining broad-spectrum resistance with good grain yields in Striga-infested fields can be recommended to rice farmers in Striga-prone areas. © 2014 The Authors. Published by Elsevier B.V. This is an open access article under the CC BY license