Transgenic expression of lactoferrin imparts enhanced resistance to head blight of wheat caused by Fusarium graminearum

<p>Abstract</p> <p>Background</p> <p>The development of plant gene transfer systems has allowed for the introgression of alien genes into plant genomes for novel disease control strategies, thus providing a mechanism for broadening the genetic resources available to plant breeders. Using the tools of plant genetic engineering, a broad-spectrum antimicrobial gene was tested for resistance against head blight caused by <it>Fusarium graminearum </it>Schwabe, a devastating disease of wheat (<it>Triticum </it><it>aestivum </it>L.) and barley (<it>Hordeum vulgare </it>L.) that reduces both grain yield and quality.</p> <p>Results</p> <p>A construct containing a bovine lactoferrin cDNA was used to transform wheat using an <it>Agrobacterium</it>-mediated DNA transfer system to express this antimicrobial protein in transgenic wheat. Transformants were analyzed by Northern and Western blots to determine lactoferrin gene expression levels and were inoculated with the head blight disease fungus <it>F</it>. <it>graminearum</it>. Transgenic wheat showed a significant reduction of disease incidence caused by <it>F. graminearum </it>compared to control wheat plants. The level of resistance in the highly susceptible wheat cultivar Bobwhite was significantly higher in transgenic plants compared to control Bobwhite and two untransformed commercial wheat cultivars, susceptible Wheaton and tolerant ND 2710. Quantification of the expressed lactoferrin protein by ELISA in transgenic wheat indicated a positive correlation between the lactoferrin gene expression levels and the levels of disease resistance.</p> <p>Conclusions</p> <p>Introgression of the lactoferrin gene into elite commercial wheat, barley and other susceptible cereals may enhance resistance to <it>F. graminearum</it>.</p

Development of broad spectrum virus resistance in plants

Viruses are widely accepted as the major class of plant pathogens responsible for significant crop losses that reduce the quality and quantity of product biomass. The objective of this study is to minimize yield losses by developing two efficient approaches based on (1) pathogen targeted resistance and (2) RNA interference (RNAi). In the first approach, the oligoadenylate synthetase (OAS), a mammalian antiviral pathway, was utilized to develop broad spectrum virus resistance in plants. It is composed of two enzymes - a 2,5A synthetase and a 2,5A-dependent RNase L. Suitability of the antiviral pathway was evaluated in transgenic tobacco (N. tabacum and N. benthamiana), soybeans and wheat plants. Tobacco plants expressing the OAS transgenes showed resistance to Tobacco mosaic virus (TMV), Tobacco etch virus (TEV) and Cucumber mosaic virus (CMV D and Y strains). Similarly, transgenic soybean carrying the OAS system was evaluated for resistance against Soybean mosaic virus (SMV) and Bean pod mosaic virus (BPMV), and transgenic wheat against Wheat streak mosaic virus (WSMV). These results showed that the system has the potential to provide broad spectrum resistance against different viruses and can confer resistance to different crops. The second approach utilizes RNA interference, a widely used method for down regulating targeted transcripts and developing disease-resistant plants. RNAi is a type of post-transcriptional gene silencing (PTGS) that relies on the presence of dsRNA to induce silencing complex leading to the degradation or inactivation of the target mRNA. A direct repeat induced gene silencing (DRIGS) system was used as a tool for developing broad spectrum virus resistance in plants. Short viral fragments from different viruses were fused in the silencing locus of the DRIGS vector and transformed into Arabidopsis, N. tabacum and N. benthamiana. Virus specific siRNAs were detected in the transgenic plants. TMV and TEV inoculated transformed plants remained symptomless and virus-free. These results show that the system simultaneously provides broad spectrum resistance to multiple viruses

Engineered plant virus resistance

Virus diseases are among the key limiting factors that cause significant yield loss and continuously threaten crop production. Resistant cultivars coupled with pesticide application are commonly used to circumvent these threats. One of the limitations of the reliance on resistant cultivars is the inevitable breakdown of resistance due to the multitude of variable virus populations. Similarly, chemical applications to control virus transmitting insect vectors are costly to the farmers, cause adverse health and environmental consequences, and often result in the emergence of resistant vector strains. Thus, exploiting strategies that provide durable and broad-spectrum resistance over diverse environments are of paramount importance

Transgenic expression of lactoferrin imparts enhanced resistance to head blight of wheat caused by \u3ci\u3eFusarium graminearum\u3c/i\u3e

Background: The development of plant gene transfer systems has allowed for the introgression of alien genes into plant genomes for novel disease control strategies, thus providing a mechanism for broadening the genetic resources available to plant breeders. Using the tools of plant genetic engineering, a broad-spectrum antimicrobial gene was tested for resistance against head blight caused by Fusarium graminearum Schwabe, a devastating disease of wheat (Triticum aestivum L.) and barley (Hordeum vulgare L.) that reduces both grain yield and quality. Results: A construct containing a bovine lactoferrin cDNA was used to transform wheat using an Agrobacterium-mediated DNA transfer system to express this antimicrobial protein in transgenic wheat. Transformants were analyzed by Northern and Western blots to determine lactoferrin gene expression levels and were inoculated with the head blight disease fungus F. graminearum. Transgenic wheat showed a significant reduction of disease incidence caused by F. graminearum compared to control wheat plants. The level of resistance in the highly susceptible wheat cultivar Bobwhite was significantly higher in transgenic plants compared to control Bobwhite and two untransformed commercial wheat cultivars, susceptible Wheaton and tolerant ND 2710. Quantification of the expressed lactoferrin protein by ELISA in transgenic wheat indicated a positive correlation between the lactoferrin gene expression levels and the levels of disease resistance. Conclusions: Introgression of the lactoferrin gene into elite commercial wheat, barley and other susceptible cereals may enhance resistance to F. graminearum

Differential regulation of miRNAs involved in the susceptible and resistance responses of wheat cultivars to wheat streak mosaic virus and Triticum mosaic virus

Abstract Background Wheat streak mosaic virus (WSMV) and Triticum mosaic virus (TriMV) are components of the wheat streak mosaic virus disease complex in the Great Plains region of the U.S.A. and elsewhere. Co-infection of wheat with WSMV and TriMV causes synergistic interaction with more severe disease symptoms compared to single infections. Plants are equipped with multiple antiviral mechanisms, of which regulation of microRNAs (miRNAs) is a potentially effective constituent. In this investigation, we have analyzed the total and relative expression of miRNA transcriptome in two wheat cultivars, Arapahoe (susceptible) and Mace (temperature-sensitive-resistant), that were mock-inoculated or inoculated with WSMV, TriMV, or both at 18 °C and 27 °C. Results Our results showed that the most abundant miRNA family among all the treatments was miRNA166, followed by 159a and 168a, although the order of the latter two changed depending on the infections. When comparing infected and control groups, twenty miRNAs showed significant upregulation, while eight miRNAs were significantly downregulated. Among them, miRNAs 9670-3p, 397-5p, and 5384-3p exhibited the most significant upregulation, whereas miRNAs 319, 9773, and 9774 were the most downregulated. The comparison of infection versus the control group for the cultivar Mace showed temperature-dependent regulation of these miRNAs. The principal component analysis confirmed that less abundant miRNAs among differentially expressed miRNAs were strongly correlated with the inoculated symptomatic wheat cultivars. Notably, miRNAs 397-5p, 398, and 9670-3p were upregulated in response to WSMV and TriMV infections, an observation not yet reported in this context. The significant upregulation of these three miRNAs was further confirmed with RT-qPCR analysis; in general, the RT-qPCR results were in agreement with our computational analysis. Target prediction analysis showed that the miRNAs standing out in our analysis targeted genes involved in defense response and regulation of transcription. Conclusion Investigation into the roles of these miRNAs and their corresponding targets holds promise for advancing our understanding of the mechanisms of virus infection and possible manipulation of these factors for developing durable virus resistance in crop plants

Additional file 1 of Differential regulation of miRNAs involved in the susceptible and resistance responses of wheat cultivars to wheat streak mosaic virus and Triticum mosaic virus

Supplementary Material 1