Comparative secretome analysis of Striga and Cuscuta species identifies candidate virulence factors for two evolutionarily independent parasitic plant lineages

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

Farming with crops and rocks to address global climate, food and soil security

The magnitude of future climate change could be moderated by immediately reducing the amount of CO 2 entering the atmosphere as a result of energy generation and by adopting strategies that actively remove CO 2 from it. Biogeochemical improvement of soils by adding crushed, fast-reacting silicate rocks to croplands is one such CO 2 -removal strategy. This approach has the potential to improve crop production, increase protection from pests and diseases, and restore soil fertility and structure. Managed croplands worldwide are already equipped for frequent rock dust additions to soils, making rapid adoption at scale feasible, and the potential benefits could generate financial incentives for widespread adoption in the agricultural sector. However, there are still obstacles to be surmounted. Audited field-scale assessments of the efficacy of CO 2 capture are urgently required together with detailed environmental monitoring. A cost-effective way to meet the rock requirements for CO 2 removal must be found, possibly involving the recycling of silicate waste materials. Finally, issues of public perception, trust and acceptance must also be addressed

A2ML1 and otitis media: novel variants, differential expression, and relevant pathways

A genetic basis for otitis media is established, however, the role of rare variants in disease etiology is largely unknown. Previously a duplication variant within A2ML1 was identified as a significant risk factor for otitis media in an indigenous Filipino population and in US children. In this report exome and Sanger sequencing was performed using DNA samples from the indigenous Filipino population, Filipino cochlear implantees, US probands, Finnish, and Pakistani families with otitis media. Sixteen novel, damaging A2ML1 variants identified in otitis media patients were rare or low-frequency in population-matched controls. In the indigenous population, both gingivitis and A2ML1 variants including the known duplication variant and the novel splice variant c.4061 + 1 G>C were independently associated with otitis media. Sequencing of salivary RNA samples from indigenous Filipinos demonstrated lower A2ML1 expression according to the carriage of A2ML1 variants. Sequencing of additional salivary RNA samples from US patients with otitis media revealed differentially expressed genes that are highly correlated with A2ML1 expression levels. In particular, RND3 is upregulated in both A2ML1 variant carriers and high-A2ML1 expressors. These findings support a role for A2ML1 in keratinocyte differentiation within the middle ear as part of otitis media pathology and the potential application of ROCK inhibition in otitis media

FUT2 Variants Confer Susceptibility to Familial Otitis Media

Non-secretor status due to homozygosity for the common FUT2 variant c.461G>A (p.Trp154∗) is associated with either risk for autoimmune diseases or protection against viral diarrhea and HIV. We determined the role of FUT2 in otitis media susceptibility by obtaining DNA samples from 609 multi-ethnic families and simplex case subjects with otitis media. Exome and Sanger sequencing, linkage analysis, and Fisher exact and transmission disequilibrium tests (TDT) were performed. The common FUT2 c.604C>T (p.Arg202∗) variant co-segregates with otitis media in a Filipino pedigree (LOD = 4.0). Additionally, a rare variant, c.412C>T (p.Arg138Cys), is associated with recurrent/chronic otitis media in European-American children (p = 1.2 × 10−5) and US trios (TDT p = 0.01). The c.461G>A (p.Trp154∗) variant was also over-transmitted in US trios (TDT p = 0.01) and was associated with shifts in middle ear microbiota composition (PERMANOVA p 20 were combined, FUT2 variants were over-transmitted in trios (TDT p = 0.001). Fut2 is transiently upregulated in mouse middle ear after inoculation with non-typeable Haemophilus influenzae. Four FUT2 variants—namely p.Ala104Val, p.Arg138Cys, p.Trp154∗, and p.Arg202∗—reduced A antigen in mutant-transfected COS-7 cells, while the nonsense variants also reduced FUT2 protein levels. Common and rare FUT2 variants confer susceptibility to otitis media, likely by modifying the middle ear microbiome through regulation of A antigen levels in epithelial cells. Our families demonstrate marked intra-familial genetic heterogeneity, suggesting that multiple combinations of common and rare variants plus environmental factors influence the individual otitis media phenotype as a complex trait

New Rice for Africa (NERICA) cultivars exhibit different levels of post-attachment resistance against the parasitic weeds Striga hermonthica and Striga asiatica

Striga hermonthica and S. asiatica are root parasitic weeds that infect the major cereal crops of sub-Saharan Africa causing severe losses in yield. The interspecific upland NEw RICe for Africa (NERICA) cultivars are popular amongst subsistence farmers, but little is known about their post-attachment resistance against Striga. Here, we evaluate the post-attachment resistance levels of the NERICA cultivars and their parents against ecotypes of S. hermonthica and S.asiatica, characterize the phenotype of the resistance mechanisms and determine the effect of Striga on host biomass. Some NERICA cultivars showed good broad-spectrum resistance against several Striga ecotypes, whereas others showed intermediate resistance or were very susceptible. The phenotype of a resistant interaction was often characterized by an inability of the parasite to penetrate the endodermis. Moreover, some parasites formed only a few connections to the host xylem, grew slowly and remained small. The most resistant NERICA cultivars were least damaged by Striga, although even a small number of parasites caused a reduction in above-ground host biomass. The elucidation of the molecular genetic basis of the resistance mechanisms and tolerance would allow the development of cultivars with multiple, durable resistance for use in farmers' fields

Tissue culture protocols for the obligate parasitic plant Striga hermonthica and implications for host-parasite co-cultivation

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

Genome-enabled discovery of candidate virulence loci in Striga hermonthica, a devastating parasite of African cereal crops

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