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Genome-enabled discovery of candidate virulence loci in Striga hermonthica, a devastating parasite of African cereal crops
New Phytologist ( IF 8.3 ) Pub Date : 2022-06-14 , DOI: 10.1111/nph.18305
Suo Qiu 1 , James M Bradley 1 , Peijun Zhang 1 , Roy Chaudhuri 1 , Mark Blaxter 2, 3 , Roger K Butlin 1, 4 , Julie D Scholes 1
Affiliation  

Introduction

Plants are constantly challenged by diverse parasites. As a consequence, they have evolved sophisticated surveillance systems to detect and protect themselves against parasite invasion (Wu et al., 2018; Kanyuka & Rudd, 2019). In turn, plant parasites have evolved suites of proteins, miRNAs, or other molecules that are delivered into host plants to facilitate colonisation (virulence factors (VFs)) (Win et al., 2012; Mitsumasu et al., 2015; Ceulemans et al., 2021; Mitchum & Liu, 2022) and they are pivotal in determining the outcome of a parasite–plant interaction.

Parasitic plants have evolved independently at least 12 times (Kuijt, 1969; Westwood et al., 2010). Regardless of evolutionary origin, parasitic plants possess a multicellular organ called the ‘haustorium’, through which direct structural and physiological connections are formed with their host plant (Westwood, 2013; Yoshida et al., 2016). This allows them to abstract water, organic and inorganic nutrients. In addition, the haustorium is increasingly recognised to play a role in host manipulation, through the movement of parasite VFs into the host plant (Shahid et al., 2018; Clarke et al., 2019). An example is provided by a particular ‘race’ of Striga gesnerioides, which delivers a small, secreted leucine-rich repeat (LRR) domain-containing effector (Suppressor of Host Resistance 4z (SHR4z)) into cowpea host cells, whereupon it triggers rapid turnover of the E3 ubiquitin ligase, VuPOB1, a positive regulator of the host's defence response (Su et al., 2020).

Striga is a genus of obligate, root-parasitic plants within the Orobanchaceae (Parker & Riches, 1993; Spallek et al., 2013). One species in particular, Striga hermonthica, infests rain-fed rice, maize, sorghum and millets, leading to devastating losses in crop yields for resource-poor farmers in sub-Saharan Africa (Scholes & Press, 2008; Rodenburg et al., 2016). Control of S. hermonthica is extremely difficult as the parasite is an obligate outbreeder, with high fecundity, wide dispersal and a persistent, long-lived seed bank (Parker & Riches, 1993) leading to a large effective population size (Huang et al., 2012). Resistant crop varieties are a crucial component of successful control strategies (Scholes & Press, 2008) however, even for crop varieties considered highly resistant, genetic variation within parasite populations is such that a few individuals can overcome host resistance and form successful attachments (Gurney et al., 2006; Cissoko et al., 2011). To develop crop varieties with durable resistance against S. hermonthica, it is vital to understand the repertoire, mode of action and genetic variability of parasite VFs (Timko et al., 2012; Rodenburg et al., 2017). Given the highly polymorphic populations of S. hermonthica and genetic diversity of the seed bank, we hypothesised that S. hermonthica is likely to possess suites of VFs that allow it to overcome layers of resistance in multiple host plant varieties. The aim of this study was to discover candidate genes encoding polymorphic VFs in S. hermonthica.

To achieve our aims we combined two complementary approaches. First, we assembled and annotated the genome of S. hermonthica, and developed a pipeline for computational prediction of putative secreted proteins (the secretome) and candidate VFs. The assembled genome was then used as a reference for an experimental, population genomics analysis, to compare DNA sequence variants in bulked (pooled) samples of S. hermonthica grown on a susceptible (NERICA-7) or resistant (NERICA-17) rice host (Fig. 1a i,ii). This allowed us to scan for loci in the S. hermonthica genome where the selection imposed by the resistant host had elevated the frequency of alleles contributing to successful colonisation (termed ‘virulence’ alleles) (Fig. 1b–d). A similar approach was used to identify candidate genomic regions associated with resistance in Solanum vernei to the potato cyst nematode, Globodera pallida (Eoche-Bosy et al., 2017). The intersection between genes encoding predicted VFs and genes with highly significant allele frequency differences in the genome scan of S. hermonthica, revealed a set of candidate virulence loci encoding proteins with many functions, for example, cell wall modification, protease or protease inhibitor and receptor-like protein kinase activities. Our results suggest that diverse strategies are used by S. hermonthica to overcome different layers of host resistance, resulting in a polygenic basis of virulence in this parasite.

Details are in the caption following the image
Fig. 1
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Experimental strategy for the identification of the Striga hermonthica virulence loci. Striga hermonthica (Kibos accession) were grown on susceptible (NERICA 7) and resistant (NERICA 17) rice hosts (a). The whole rice root systems show many S. hermonthica individuals parasitising the roots of NERICA 7 (i) whilst only two individuals (red circles) were able to overcome the resistance response of NERICA 17 (ii). Transverse sections show S. hermonthica invading rice roots for a representative susceptible (iii) and resistant (iv–vi) interaction 7 d after inoculation. In the successful host–parasite interaction parasite intrusive cells (PIC) have breached the endodermis and have made connections with the host's xylem (iii). In the resistant rice variety several phenotypes are observed; The parasite invades the host root cortex but is unable to penetrate the suberised endodermis (iv, v); the parasite penetrates the endodermis but is unable to form connections with the host xylem (vi). H, host root; P, parasite. Bars, 5 μm. Our experimental strategy was based on the prediction that many S. hermonthica genotypes would grow on NERICA 7 but only highly virulent genotypes would grow on NERICA 17 (b). Samples of 100 S. hermonthica plants were bulked to generate three sequencing pools from each host variety (c). We expected that background loci would not differ in allele frequency between pools, but virulence alleles (and neutral alleles in linkage disequilibrium) would have increased frequency in all pools from the resistant host, allowing us to identify candidate loci (d). S1–S3, sequencing pools from susceptible plants (NERICA-7); R1–R3, sequencing pools from resistant plants (NERICA-17).


中文翻译:

通过基因组发现 Striga hermonthica 的候选毒力位点,这是一种非洲谷类作物的毁灭性寄生虫

介绍

植物不断受到各种寄生虫的挑战。因此,它们进化出复杂的监视系统来检测和保护自己免受寄生虫入侵(Wu等人2018 年;Kanyuka 和 Rudd,  2019 年)。反过来,植物寄生虫已经进化出一系列蛋白质、miRNA 或其他分子,这些分子被输送到宿主植物中以促进定殖(毒力因子 (VF))(Win等人,  2012 年;Mitsumasu等人,  2015 年;Ceulemans等人.,  2021;Mitchum & Liu,  2022 ),它们在决定寄生虫与植物相互作用的结果方面起着关键作用。

寄生植物至少独立进化了 12 次 (Kuijt,  1969 ; Westwood et al .,  2010 )。无论进化起源如何,寄生植物都拥有一个称为“吸器”的多细胞器官,通过该器官与其寄主植物形成直接的结构和生理联系(Westwood,2013 年;Yoshida等人,  2016 年)。这使它们能够提取水、有机和无机营养物。此外,越来越多的人认识到吸器通过将寄生虫 VF 移动到宿主植物中而在宿主操纵中发挥作用(Shahid等人,  2018 年;Clarke等人, 2019 年)。Striga gesnerioides的一个特定“种族”提供了一个例子,它将一个小的、分泌的富含亮氨酸重复 (LRR) 结构域的包含效应子(宿主抗性抑制因子 4z (SHR4z))传递到豇豆宿主细胞中,随后它触发快速E3 泛素连接酶 VuPOB1 的转换,它是宿主防御反应的正调节因子 (Su et al .,  2020 )。

独脚金是列当科中专性根寄生植物的一个属(Parker & Riches,  1993 年;Spallek等人,  2013 年)。尤其是Striga hermonthica 一种,它侵染了雨养水稻、玉米、高粱和小米,导致撒哈拉以南非洲资源贫乏的农民的作物产量遭受毁灭性损失(Scholes & Press,  2008 年;Rodenburg等人,  2016 年) ). 控制S. hermonthica极其困难,因为该寄生虫是专性远系繁殖者,具有高繁殖力、广泛传播和持久、长寿命的种子库(Parker & Riches,  1993) 导致有效种群规模较大 (Huang et al .,  2012 )。抗性作物品种是成功控制策略的重要组成部分(Scholes & Press,  2008)然而,即使对于被认为具有高度抗性的作物品种,寄生虫种群内的遗传变异也是如此,一些个体可以克服宿主抗性并形成成功的依附(Gurney等等人,  2006 年;Cissoko等人,  2011 年)。要开发对S. hermonthica具有持久抗性的作物品种,了解寄生虫 VF 的所有组成部分、作用方式和遗传变异性至关重要(Timko等人, 2012 年;罗登堡等人,  2017 年)。鉴于S. hermonthica的高度多态性种群和种子库的遗传多样性,我们假设S. hermonthica可能拥有一系列 VF,使其能够克服多种寄主植物品种的抗性层。本研究的目的是在S. hermonthica中发现编码多态性 VF 的候选基因。

为了实现我们的目标,我们结合了两种互补的方法。首先,我们组装并注释了S. hermonthica的基因组,并开发了用于计算预测假定分泌蛋白(分泌蛋白组)和候选 VF 的管道。然后将组装的基因组用作实验性群体基因组学分析的参考,以比较在易感(NERICA-7)或抗性(NERICA-17)水稻宿主上生长的S. hermonthica的大量(合并)样本中的 DNA 序列变异(图 1a i,ii)。这使我们能够扫描S. hermonthica中的位点抗性宿主施加的选择提高了有助于成功定植的等位基因频率的基因组(称为“毒力”等位基因)(图1b-d)。一种类似的方法被用于鉴定与马铃薯孢囊线虫Globodera pallida的抗性相关的候选基因组区域( Eoche -Bosy等人,  2017 年)。在S. hermonthica的基因组扫描中编码预测 VF 的基因与具有高度显着等位基因频率差异的基因之间的交集, 揭示了一组候选毒力位点编码具有许多功能的蛋白质,例如,细胞壁修饰、蛋白酶或蛋白酶抑制剂和受体样蛋白激酶活性。我们的结果表明,S. hermonthica使用不同的策略来克服不同层的宿主抗性,从而导致这种寄生虫的毒力的多基因基础。

详细信息在图片后面的标题中
图。1
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鉴定Striga hermonthica毒力位点的实验策略。Striga hermonthica(Kibos 登录号)在易感(NERICA 7)和抗性(NERICA 17)水稻宿主(a)上生长。整个水稻根系显示许多S. hermonthica个体寄生在 NERICA 7 (i) 的根部,而只有两个个体(红色圆圈)能够克服 NERICA 17 (ii) 的抗性反应。横截面显示S. hermonthica接种后 7 天侵入水稻根系的代表性易感 (iii) 和抗性 (iv-vi) 相互作用。在成功的宿主-寄生虫相互作用中,寄生虫侵入细胞 (PIC) 已经突破了内皮层并与宿主的木质部建立了联系 (iii)。在抗性水稻品种中观察到几种表型;寄生虫侵入宿主根皮层,但无法穿透栓化的内皮层 (iv, v);寄生虫穿透内皮但无法与宿主木质部 (vi) 形成连接。H,主机根;P,寄生虫。条形,5 微米。我们的实验策略基于以下预测:许多S. hermonthica基因型会在 NERICA 7 上生长,但只有高毒性基因型会在 NERICA 17 (b) 上生长。100 个S. hermonthica的样本植物被批量化以从每个宿主品种 (c) 生成三个测序库。我们预计背景基因座在池之间的等位基因频率上不会不同,但毒力等位基因(和连锁不平衡中的中性等位基因)会增加来自抗性宿主的所有池中的频率,从而使我们能够识别候选基因座(d)。S1–S3,来自易感植物的测序池 (NERICA-7);R1–R3,来自抗性植物 (NERICA-17) 的测序库。
更新日期:2022-06-14
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