Integrated Transcriptome and Proteome Analysis Reveals That Cell Wall Activity Affects Phelipanche aegyptiaca Parasitism.

Phelipanche aegyptiaca can infect many crops, causing large agricultural production losses. It is important to study the parasitism mechanism of P. aegyptiaca to control its harm. In this experiment, the P. aegyptiaca HY13M and TE9M from Tacheng Prefecture and Hami City in Xinjiang, respectively, were used to analyze the parasitical mechanism of P. aegyptiaca by means of transcriptome and proteome analyses. The parasitic capacity of TE9M was significantly stronger than that of HY13M in Citrullus lanatus. The results showed that the DEGs and DEPs were prominently enriched in the cell wall metabolism pathways, including "cell wall organization or biogenesis", "cell wall organization", and "cell wall". Moreover, the functions of the pectinesterase enzyme gene (TR138070_c0_g), which is involved in the cell wall metabolism of P. aegyptiaca in its parasitism, were studied by means HIGS. The number and weight of P. aegyptiaca were significantly reduced when TR138070_c0_g1, which encodes a cell-wall-degrading protease, was silenced, indicating that it positively regulates P. aegyptiaca parasitism. Thus, these results suggest that the cell wall metabolism pathway is involved in P. aegyptiaca differentiation of the parasitic ability and that the TR138070_c0_g1 gene plays an important role in P. aegyptiaca's parasitism.

Identification and genetic diversity analysis of broomrape in Xinjiang, China.

BACKGROUND: As a holoparasitic weed, broomrape has seriously threatened the production of economically important crops, such as melon, watermelon, processed tomato, and sunflower, in Xinjiang in recent years. However, the distribution and genetic diversity of broomrape populations in Xinjiang are not clear at present, which hinders their prevention and control. The purpose of this study was to identify the main species and the genetic differentiation structure of the broomrape population in Xinjiang. METHODS AND RESULTS: In the present study, 93 samples from different geographic regions of Xinjiang were collected to identify the species based on ITS and plastid rps2 regions, and the samples were also used to analyze the genetic diversity based on ISSR markers. The results showed that broomrape is not monophyletic in Xinjiang and consists of two major clades (Orobanche cf. aegyptiaca and O. cernua) and three subclades (O. cf. aegyptiaca var. tch, O. cf. aegyptiaca var. klz, and O. cernua.var. alt) based on phylogenetic analysis. Furthermore, the results of the genetic diversity analysis indicated that the average polymorphic information content and marker index were high values of 0.58 and 7.38, respectively, showing the efficiency of the ISSR markers in detecting polymorphism among the broomrape population studied. Additionally, the 11 selected primers produced 154 repeatable polymorphic bands, of which 150 were polymorphic. The genetic diversity of the samples was 37.19% within populations and 62.81% among the populations, indicating that the main genetic differentiation occurred among the populations. There was less gene exchange between populations, with a gene flow index (Nm) of 0.2961 (< 1). The UPGMA dendrogram indicated that most populations with similar geographical conditions and hosts were clustered first, and then all samples were separated into two major groups and seven subclusters. CONCLUSION: The broomrapes are mainly O. cf. aegyptiaca and O. cernua in Xinjiang, which were separated into two major groups and seven subclusters based on ISSR markers. Our results provide a theoretical basis for breeding broomrape-resistant varieties.

Comprehensive genomic characterisation of the NAC transcription factor family and its response to drought stress in Eucommia ulmoides.

The NAC transcription factor family enhances plant adaptation to environmental challenges by participating in signalling pathways triggered by abiotic stressors and hormonal cues. We identified 69 NAC genes in the Eucommia ulmoides genome and renamed them according to their chromosomal distribution. These EuNAC proteins were clustered into 13 sub-families and distributed on 16 chromosomes and 2 scaffolds. The gene structures suggested that the number of exons varied from two to eight among these EuNACs, with a multitude of them containing three exons. Duplicated events resulted in a large gene family; 12 and four pairs of EuNACs were the result of segmental and tandem duplicates, respectively. The drought-stress response pattern of 12 putative EuNACs was observed under drought treatment, revealing that these EuNACs could play crucial roles in mitigating the effects of drought stress responses and serve as promising candidate genes for genetic engineering aimed at enhancing the drought stress tolerance of E. ulmoides. This study provides insight into the evolution, diversity, and characterisation of NAC genes in E. ulmoides and will be helpful for future characterisation of putative EuNACs associated with water deficit.

First Report of Karelinia caspia (Pall.) Less as a new host of Orobanche cumana Wallr. in Xinjiang, China.

Sunflower broomrape (Orobanche cumana Wallr.) is a holoparasitic plant species which mainly parasitizes a few species of the Asteraceae in the wild and is exclusively found growing on sunflower in agricultural fields (Fernández-Martínez et al. 2015). O. cumana is a serious threat to sunflower production in Xinjiang and Inner Mongolia (Shi et al. 2015). Karelinia caspia (Pall.) Less. (Asteraceae) is an ecologically important plant species occurring across the desert ecosystems of Russia, Central Asia, and northwest China. It plays an important role in reducing wind erosion and desertification (Xu et al. 2018). During the 2018 and 2019 growing seasons, sunflower broomrape was observed parasitizing K. caspia in non-cultivated areas adjacent to sunflower fields near Beitun city (87°51'E, 47°15'N) in Xinjiang, China. Sunflower broomrape plants were identified morphologically as O. cumana according to Pujadas-Salvà and Velasco (2000). The host plants were identified morphologically as K. caspia according to Lin et al (1979). The ribosomal DNA internal transcribed spacer (ITS) and the trnL-F region of the parasite were amplified by PCR using primer pairs ITS1/ITS4 and trnL-FF/trnL-FR, respectively (Taberlet et al. 1991; Anderson et al. 2004). The ITS sequence of the parasite (Accession No. MT795725.1) showed 100% identity (675bp out of 689bp) to that of O. cernua var. cumana (KC811228.1). The trnl-F sequence of the parasite (Accession No. ON843707) showed 98% identity (675 of 689 bp) to O.cernua var. cumana (KT387722.1). Multi-locus phylogenetic analysis of the two sequences showed clustering with sunflower broomrape. The ITS region of the parasite and host was were amplified by PCR using the primer pair ITS1F/ITS4R (Taberlet et al.1991), and the ITS sequences of the host (Accession No. MT791995.1) showed 99.86% identity (728bp of 802bp) to that of K. caspia (LN607483.1). Rhizotron and pot experiments were carried out to assess the parasitic relationship between O. cumana and K. caspia. In the rhizotron experiment, 2-week-old seedlings of K. caspia were inoculated with sterilized 400 O. cumana seeds in a 15-cm petri dish filled with a sponge overlaid with glass fiber filter paper. The parasitic state of O. cumana was observed 9 days after inoculation. In another trial, seeds of K. caspia were sowed in 2-L and 4-L pots containing sand-vermiculite-compost (1:1:1 v:v:v). These pots were artificially inoculated with 50 mg of O. cumana seeds per 1 kg of substrate. After 20 and 70 days, corresponding to the early parasitic and flowering stages, respectively, of O. cumana, K. caspia plants were uprooted from the media and washed carefully. The parasitic relationship was confirmed by the attachment position of the broomrape to the K. caspia root. To our knowledge, this is the first report of O. cumana parasitizing K. caspia in Xinjiang, China. This phenomenon means that sunflower broomrape can raise up seed on a newly recognized host. Weed eradication in and near sunflower fields is a key measure to control sunflower broomrape.

Detection of Secreted Effector Proteins from Phelipanche aegyptiaca During Invasion of Melon Roots.

Parasites can interact with their host plants through the induction and delivery of secreted effector proteins that facilitate plant colonization by decomposing plant cell walls and inhibiting plant immune response to weaken the defense ability of the host. Yet effectors mediating parasitic plant-host interactions are poorly understood. Phelipanche aegyptiaca is an obligate root parasite plant causing severe yield and economic losses in agricultural fields worldwide. Host resistance against P. aegyptiaca occurred during the attachment period of parasitism. Comparative transcriptomics was used to assess resistant and susceptible interactions simultaneously between P. aegyptiaca and two contrasting melon cultivars. In total, 2,740 secreted proteins from P. aegyptiaca were identified here. Combined with transcriptome profiling, 209 candidate secreted effector proteins (CSEPs) were predicted, with functional annotations such as cell wall degrading enzymes, protease inhibitors, transferases, kinases, and elicitor proteins. A heterogeneous expression system in Nicotiana benthamiana was used to investigate the functions of 20 putatively effector genes among the CSEPs. Cluster 15140.0 can suppress BAX-triggered programmed cell death in N. benthamiana. These findings showed that the prediction of P. aegyptiaca effector proteins based on transcriptomic analysis and multiple bioinformatics software is effective and more accurate, providing insights into understanding the essential molecular nature of effectors and laying the foundation of revealing the parasite mechanism of P. aegyptiaca, which is helpful in understanding parasite-host plant interaction.

First Report of Orobanche cumana on Coleus in Xinjiang, China.

Coleus (Plectranthus scutellarioides [L.] R.Br., [syn.: Solenostemon scutellarioides], Lamiaceae) is a popular ornamental plant for its colorful and showy foliage, and widely planted as a garden plant, and a medicinal herb in some countries, including India, Indonesia, Mexico (Zhu et al. 2015). In March 2022, parasitism of broomrape, on coleus plants was found in a greenhouse (86° 3' 36" E, 44° 18' 36" N, 500 m elevation) at Shihezi University, Xinjiang, China. A few plants (6%) were parasitized with 2.5 emerged broomrape shoots per host plant. The host-parasite connection was confirmed by microscopy. Morphological characteristics of the host were consistent with coleus described by Cao et al. (2023). The broomrapes were: stem simple and slender, slightly bulbous at the base, glandular-pubescent; inflorescence usually many-flowered, lax, dense in the upper third; bracts 8 to 10 mm long, ovate-lanceolate; calyx segments free, entire, seldom bifid with markedly unequal subulate teeth; corolla markedly curvate, dorsal line inflected, white at the base, bluish violet in the upper part; stamens adaxial with filaments 6 to 7 mm long; abaxial stamens with filaments 7 to 10 mm long; gynoecium 7 to 10 mm long; ovary 4 to 5 mm long, glabrous; style with short glandular hairs; stigma white, keyed to sunflower broomrape (Orobanche cumana Wallr.) (Pujadas-Salvà and Velasco 2000). Total genomic DNA of this parasite flowers was extracted and the trnL-F gene and ribosomal DNA internal transcribed spacer (ITS) region were amplified using the primer pairs C/F and ITS1/ITS4, respectively (Taberlet et al. 1991; Anderson et al. 2004). Sequences of ITS (655 bp) and trnL-F (901 bp) were obtained (GenBank ON491818 and ON843707). BLAST analysis showed the ITS sequence was identical to that of sunflower broomrape (MK567978.1), also the trnL-F sequence matched that of sunflower broomrape (MW809408.1, identity 100%). Multi-locus phylogenetic analyses of the two sequences showed this parasite is clustered with sunflower broomrape. Together, morphological and molecular evidences confirmed the parasite on coleus plants was sunflower broomrape, a root holoparasitic plant with a narrow host range, which mainly posed a devastating threat to sunflower planting industry (Fernández-Martínez et al. 2015). To verify that coleus sunflower broomrape parasitic association, seedlings of this host were planted in 1.5-L pots containing compost-vermiculite-sand mixture (1:1:1 v:v:v) and sunflower broomrape seeds (50 mg seeds per 1 kg, soil). Three coleus seedlings, transplanted into pots without sunflower broomrape seeds, served as control. Ninety-six days later, the infected plants were smaller, their leaf color was observed to a lighter green than those of control plants and were similar to the broomrape-infected coleus plants observed in the greenhouse. The coleus roots with sunflower broomrape were carefully washed with running water, 10 to 15 emerged broomrape shoots and 14 to 22 underground attachments were observed on the coleus roots. The parasite grew well in coleus roots, from germination, attachment to host roots, and tubercles development. At the tubercle stage, the endophyte of sunflower broomrape had connected with the vascular bundle of the coleus root, confirming the sunflower broomrape-coleus connection. To the best of our knowledge, this was the first report of sunflower broomrape parasitizing coleus in Xinjiang, China. This indicates that sunflower broomrape can be propagated and survived by coleus, in fields or greenhouses with sunflower broomrape. To limit the spread of sunflower broomrape, preventive field management is needed for the coleus farmlands and greenhouse where the root holoparasite is prevalent.

Phenotypic and histological analyses on the resistance of melon to Phelipanche aegyptiaca.

Melon (Cucumis melo L.) is an economically important crop in Xinjiang, China, but its production is constrained by the parasitic plant Phelipanche aegyptiaca that attaches to the roots of many crops and causes severe stunting and loss of yield. Rhizotron, pot, and field experiments were employed to evaluate the resistance of 27 melon cultivars to P. aegyptiaca. Then, the resistant and susceptible cultivars were inoculated with P. aegyptiaca from six populations to assess their resistance stability and broad spectrum. Further microscopic and histological analyses were used to clarify the resistance phenotypes and histological structure. The results showed that Huangpi 9818 and KR1326 were more resistant to P. aegyptiaca compared to other cultivars in the rhizotron, pot, and field experiments. In addition, compared to the susceptible cultivar K1076, Huangpi 9818 and KR1326 showed broad-spectrum resistance to six P. aegyptiaca populations. These two resistant cultivars had lower P. aegyptiaca biomass and fewer and smaller P. aegyptiaca attachments on their roots compared to susceptible cultivar K1076. KR1326 (resistant) and K1076 (susceptible) were selected to further study resistance phenotypes and mechanisms. Germination-inducing activity of root exudates and microscopic analysis showed that the resistance in KR1326 was not related to low induction of P. aegyptiaca germination. The tubercles of parasite on KR1326 were observed slightly brown at 14 days after inoculation (DAI), the necrosis and arrest of parasite development occurred at 23 DAI. Histological analysis of necrosis tubercles showed that the endophyte of parasite had reached host central cylinder, connected with host xylem, and accumulation of secretions and callose were detected in neighbouring cells. We concluded that KR1326 is an important melon cultivar for P. aegyptiaca resistance that could be used to expand the genetic basis of cultivated muskmelon for resistance to the parasite.

First Report of Phelipanche aegyptiaca on Plectranthus scutellarioides in Xinjiang, China.

Coleus (Plectranthus scutellarioides [L.] R.Br.[syn.: Solenostemon scutellarioides]) is a perennial plant in the Lamiaceae family. It produces variegated leaves of various colors. It is commonly cultivated as an ornamental plant or grown in commercial greenhouses (Garibaldi et al. 2019). Phelipanche aegyptiaca Pers. is a dicotyledonous holoparasitic flowering plant that parasitizes more than 30 food crops (e.g., tomato, sunflower, and chickpea), ornamental crops, and others in different parts of the world, causing heavy economic losses (Nosratti et al. 2020). In 2016 and 2017, broomrape was observed parasitizing coleus in the greenhouse (86° 3' 36" E, 44° 18' 36" N, 500 m elevation) in Shihezi, Xinjiang, China (Supplementary Figure 1A-D). A single coleus plant could be parasitized by average 6-10 broomrape plants, and 20% of coleus plants were infested. The infection was confirmed by verifying the attachment of the broomrape to the coleus root. The inflorescences of the broomrape were normal and healthy and produced germinable seeds (germination rate: 80-90%). The morphological characteristics of the coleus are shown in Supplementary Figures 6 and 7. The main botanical features of the broomrape are as follows: (i) stem 20.65±7.07 cm tall, erect, branched, frail, rather hairy, bulbous at the base with secondary roots; (ii) inflorescence usually many-flowered, lax and cylindrical; (iii) bracts 6.87±0.93 mm long, ovate to lanceolate; (iv) calyx 1.09±0.09 cm long, shortly campanulate; (v) corolla 3.38±0.19 cm long, erect to suberect, white at the base, blue-purple in the upper part, sparsely glandular-villous; (vi) stamens 4, filaments inserted 5-6 mm from the base of the corolla, 1.26±0.11 cm long, anthers with villous; (vii) pistil 2.9±0.15 cm long, ovary glabrous, style with short glandular hairs, stigma bilobed, white (Supplementary Figure 2) (Teimoury et al. 2012; Piwowarczyk et al. 2019). For molecular identification, total genomic DNA was extracted from the flowers of the broomrape (found parasitizing coleus plants), and the ribosomal protein S2 (rps2) and ribosomal DNA internal transcribed spacer (ITS) region were amplified by PCR using the primer pairs rps2F/rps2R, ITS1/ITS4 (Table 1) (Park et al. 2007; Anderson et al. 2004). Two sequences with 580 bp (ITS) and 443 bp (rps2) were obtained (GenBank accession No. MW811482 and MW883573). BLAST analysis showed that the ITS sequence was most similar (identity 100%) to P. aegyptiaca (KC811171) and the rps2 sequence (identity 99%) also matched that of P. aegyptiaca (KC814957). Phylogenetic analysis of the ITS regions and rps2 genes showed that broomrape was fallen into P. aegyptiaca groups (Supplementary Figure 3). Morphological and molecular findings strongly support the conclusion that the broomrape on coleus was P. aegyptiaca. In order to verify that coleus was a host of P. aegyptiaca, coleus seedlings were collected and moved to 1.5-L pots containing a mixture of compost-vermiculite-sand (1:1:1 v:v:v) and seeds of P. aegyptiaca harvested from the host coleus (50 mg of P. aegyptiaca seeds per 1 kg of the substrate). Another three coleus seedlings were transplanted into pots of the same size containing the same mixture as above without P. aegyptiaca seeds. These served as controls. After 90 days of inoculation, the leaves of the infected hosts were lighter in color than those of uninfected hosts (Supplementary Figures 4A, 6). The roots of coleus and P. aegyptiaca were carefully washed with water, and an average of 3-4 emerged broomrape shoots and 50-60 underground attachments were observed on coleus roots (Supplementary Figure 4B). P. aegyptiaca can develop normally in the root of the coleus plant, from germination through attachment to host roots and development of tubercles (Supplementary Figure 5 A-E). Longitudinal and transverse sections of the parasite and host roots at the tubercle stage revealed that the endophytic tissues of P. aegyptiaca had reached and connected to the host vascular bundle (Supplementary Figure 5F-I), confirming the normal biological development and function of P. aegyptiaca haustoria. To the best of our knowledge, this is the first report of P. aegyptiaca parasitizing coleus in Xinjiang, China. Coleus is a very widely cultivated horticultural ornamental plant, and it grows in the same environments favored by P. aegyptiaca; so, the plant can aid the transmission of P. aegyptiaca to previously clear regions. It is necessary to improve the management of coleus in places where P. aegyptiaca is prevalent so as to reduce its spread. References: Garibaldi, A., et al. 2019. Plant Dis. 104:590. https://doi.org/10.1094/PDIS-07-19-1399-PDN Crossref, ISI, Google Scholar Nosratti, I., et al. 2020. Weed Sci. 68:555-564. https://doi.org/10.1017/wsc.2020.61 Crossref, ISI, Google Scholar Teimoury, M., et al. 2012. Plant Dis. 96:1232. https://doi.org/10.1094/PDIS-01-12-0068-PDN Crossref, ISI, Google Scholar Piwowarczyk, R., et al. 2019. Phytotaxa. 386:001-106. https://doi.org/10.11646/phytotaxa.386.1.1 Crossref, ISI, Google Scholar Park, J. M., et al. 2007. Mol. Phylogenet. Evol. 43: 974-985. https://doi.org/10.1016/j.ympev.2006.10.011 Crossref, ISI, Google Scholar Anderson, I. C., et al. 2004. Environ. Microbiol. 6: 769-779. https://doi.org/10.1111/j.1462-2920.2004.00675.x Crossref, ISI, Google Scholar.

Identification of risk areas for Orobanche cumana and Phelipanche aegyptiaca in China, based on the major host plant and CMIP6 climate scenarios.

Parasitic broomrape of the genus Orobanche poses a formidable threat to producing many crops in Europe, Africa, and Asia. Orobanche cumana and Phelipanche aegyptiaca are two of China's most destructive root parasitic plants, causing extreme sunflower, tomato, melon, and tobacco damage. However, the potentially suitable areas of O. cumana and P. aegyptiaca in China have not been predicted, and little is known about the important environmental factors that affect their extension. Due to their invasiveness and economic importance, studying how climate change and host plants may affect broomrapes' distribution is necessary. In the study, we first predicted the potentially suitable areas of the invasive weeds (O. cumana and P. aegyptiaca) and their susceptible host plants (Helianthus annuus and Solanum lycopersicon) using MaxEnt. Then, the risk zones and distribution shifts of two broomrapes under different climate conditions were identified by incorporating the distribution of their susceptible host plants. The results highlighted that the potential middle- and high-risk zones for O. cumana and P. aegyptiaca amounted to 197.88 × 104 km2 and 12.90 × 104 km2, respectively. Notably, Xinjiang and Inner Mongolia were the highest-risk areas within the distribution and establishment of O. cumana and P. aegyptiaca. Elevation and topsoil pH were the decisive factors for shaping O. cumana distribution; precipitation seasonality and annual precipitation were the dominant bioclimatic variables limiting the spread of P. aegyptiaca. The potentially suitable areas and risk zones of O. cumana would decrease significantly, and those of P. aegyptiaca would fluctuate slightly under future climate change scenarios. Overall, our study suggested that the two broomrapes' risk zones will significantly northward to higher latitudes. The results will provide suggestions for preventing O. cumana and P. aegyptiaca.

Lipopeptide production by Bacillus atrophaeus strain B44 and its biocontrol efficacy against cotton rhizoctoniosis.

OBJECTIVES: An assay was conducted to show the comparisons the effects of nine metal ions on antagonistic metabolites (lipopeptides, siderophores and gibberellins) by Bacillus atrophaeus strain B44 using well-diffusion assays, matrix-assisted laser desorption ionization time-of-flight mass spectrometry (MALDI-TOF-MS) analysis, chrome azurol S plus mannitol salt agar (CAS-MSA) tests, and reversed-phase high-performance liquid chromatography (RP-HPLC) analysis. This assay is also designed to demonstrate the biocontrol efficacy of B44 against cotton rhizoctoniosis using pot culture tests. RESULTS: Both the lipopeptide yield and the antimicrobial activity of B44 increase with the MnSO4, MgSO4, CaCO3, and CuSO4 treatments and either have no effect or decreased lipopeptide yield and antimicrobial activity with the FeSO4, K2HPO4, KCl, KH2PO4 and ZnSO4 treatments. The medium containing MgSO4 has no significant effect on either the lipopeptide yield or antimicrobial activity. MALDI-TOF-MS analysis shows a broad range of m/z peaks, indicating that strain B44 produces a complex mixture of iturin, surfactin, and fengycin lipopeptides. Gibberellin production by strain B44 varies greatly depending on the culture medium, and the siderophore production is not significantly affected by the culture medium. Pot tests show that lipopeptide production affects the disease control efficacy of strain B44. CONCLUSION: The biocontrol efficacy of B. atrophaeus strain B44 is related to the lipopeptide yield. Moreover, B. atrophaeus strain B44 significantly increases the size of cotton seedlings, which is related to the GA3 concentration.

Development of droplet digital PCR for the detection of Tilletia laevis, which causes common bunt of wheat, based on the SCAR marker derived from ISSR and real-time PCR.

Common bunt of wheat caused by Tilletia laevis and/or T. caries (syn. T. tritici), is a major disease in wheat-growing regions worldwide that could lead to 80% or even total loss of production. Even though T. laevis can be distinguished from T. caries on the bases of morphology of teliospores using microscopy technique. However, molecular methods could serve as an additional method to quantify the pathogen. To develop a rapid diagnostic and quantify method, we employed the ISSR molecular marker for T. laevis in this study. The primer ISSR857 generated a polymorphic pattern displaying a 1385 bp T. laevis-specific DNA fragment. A pair of specific primers (L57F/L57R) was designed to amplify a sequence-characterized amplified region (SCAR) (763 bp) for the PCR detection assay. The primers amplified the DNA fragment in the tested isolates of T. laevis but failed in the related species, including T. caries. The detection limit of the primer set (L57F/L57R) was 5 ng/µl of DNA extracted from T. laevis teliospores. A SYBR Green I real-time PCR method for detecting T. laevis with a 100 fg/µl detection limit and droplet digital PCR with a high sensitivity (30 fg/µl detection limit) were developed; this technique showed the most sensitive detection compared to the SCAR marker and SYBR Green I real-time PCR. Additionally, this is the first study related the detection of T. laevis with the droplet digital PCR method.

Development of ISSR-derived SCAR Marker and SYBR Green I Real-time PCR Method for Detection of Teliospores of Tilletia laevis Kühn.

Common bunt, caused by Tilletia laevis Kühn [syn. T. foetida (Wallr) Liro] and Tilletia tritici (Bjerk.) Wint. [syn. T. caries (DC) Tul.], is an important wheat disease worldwide. To quickly differentiate the closely related fungi T. laevis, T. tritici and Tilletia controversa (a pathogen that causes dwarf bunt of wheat and has been requested as a quarantined pathogen in many countries), a rapid diagnostic and detection method for an ISSR molecular marker was developed for the first time in this study. Based on the T. laevis-specific band (1300 bp) amplified by the primer ISSR860, a pair of SCAR primers (L60F/L60R) was designed to amplify a specific 660-bp DNA fragment from the isolates of T. laevis but not other related pathogens. The detection limit of the SCAR marker was 0.4 ng/µl of DNA from T. laevis; moreover, a SYBR Green I real-time PCR method was also successfully developed based on the SCAR marker with the detection limit of 10 fg/µl T. laevis DNA. This is the first report of a rapid, specific and highly sensitive SCAR marker and SYBR Green I real-time PCR method for detection of the teliospores of T. laevis based on ISSR technology. This method allows highly efficient, rapid and accurate differentiation of the pathogen from related pathogens, especially from the very similar pathogens T. tritici and T. controversa.

Global Transcriptomic Analysis Reveals the Mechanism of Phelipanche aegyptiaca Seed Germination.

Phelipanche aegyptiaca is one of the most destructive root parasitic plants of Orobanchaceae. This plant has significant impacts on crop yields worldwide. Conditioned and host root stimulants, in particular, strigolactones, are needed for unique seed germination. However, no extensive study on this phenomenon has been conducted because of insufficient genomic information. Deep RNA sequencing, including de novo assembly and functional annotation was performed on P. aegyptiaca germinating seeds. The assembled transcriptome was used to analyze transcriptional dynamics during seed germination. Key gene categories involved were identified. A total of 274,964 transcripts were determined, and 53,921 unigenes were annotated according to the NR, GO, COG, KOG, and KEGG databases. Overall, 5324 differentially expressed genes among dormant, conditioned, and GR24-treated seeds were identified. GO and KEGG enrichment analyses demonstrated numerous DEGs related to DNA, RNA, and protein repair and biosynthesis, as well as carbohydrate and energy metabolism. Moreover, ABA and ethylene were found to play important roles in this process. GR24 application resulted in dramatic changes in ABA and ethylene-associated genes. Fluridone, a carotenoid biosynthesis inhibitor, alone could induce P. aegyptiaca seed germination. In addition, conditioning was probably not the indispensable stage for P. aegyptiaca, because the transcript level variation of MAX2 and KAI2 genes (relate to strigolactone signaling) was not up-regulated by conditioning treatment.