bluuebunny/crossref_metadata_2025 huggingface dataset:
Sub-Specific Populations of Verticillium dahliae and their Roles in Vascular Wilt Pathogsystems
Authors: Rowe Randall C. , Katan Jaacov , Katan Talma , Tsror Leah
Published: December 2021

Verticillium dahliae is an economically important pathogen causing vascular wilt on over 160 plant species. In North America, potato early dying is a significant disease of potato, especially in the midwest and Pacific northwest states. This disease is caused by the fungus Verticillium dahliae and in some cases involves a synergistic interaction with root-lesion nematodes, primarily Pratylenchus penetrans. In Israel, Verticillium wilt occurs in many regions and inflicts serious losses in potato, cotton, and other crops. Objectives of this project were to establish a large collection of isolates of Verticillium dahliae from potato (USA) and several host plants (Israel) and to characterize and compare the isolates with regard to morphology, vegetative compatibility group (VCG), and pathogenic capabilities on several hosts. Isolations were made from 224 commercial lots of certified potato seed tubers from across N. America and 87 potato fields located in the Columbia Basin of Oregon and Washington. A large collection of isolates from central U.S. states already existed. In Israel, 47 field sites were sampled and isolates of Verticillium dahliae were recovered from 13 host plant species and from soil. Potato isolates from N. America were tested for vegetative compatibility and all found to be in VCG 4 with about 2/3 in VCG 4A and the rest in VCG 4B. VCG 4A isolates were significantly more aggressive on potato than VCG 4B isolates and were more likely to interact synergistically with P. penetrans. The Israeli isolates fell into three vegetative compatibility groups. Nearly all (> 90%) VCG2B and VCG 4B isolates were recovered from the northern and southern parts of Israel, respectively, with some overlap in central areas. Several pathotypes were defined in cotton, using cotton and eggplant together as differentials. All VCG 2B isolates from cotton caused severe disease in cotton, while VCG 2A and VCG 4B isolates from several crops were much less aggressive to cotton. When Israeli isolates of VCGs 2A, 2B and 4B were inoculated to potato and tomato, VCG 4B isolates caused much more severe disease on potato and VCG 2A isolates caused much more severe disease in tomato. Differential patterns of pathogenicity and aggressiveness of these VCGs on potato and tomato were consistent regardless of the host plant of origin. Isolates of the same VCG resembled one another more than isolates from different VCGs based on colony and microsclerotial morphology, temperature responses and, partially, in pathogenicity. Vegetative compatibility grouping of V. dahliae in Israel appears closely associated with specific pathogenicity and other phenotypic traits. The absence of VCG 4A in Israel is significant. VCG patterns among Verficillium populations are useful to predict relatedness and pathogenic potential in both countries.
Enhancement of Essential Amino Acids in Cereal Seeds: Four Approaches to Increased Lysine Content
Authors: Anderson Olin D. , Galili Gad , Blechl Ann E.
Published: December 2021

Cereal seeds are the basis of the human diet, and their amino acid composition is thus of major nutritional and economic importance. Currently, deficiencies in essential amino acids are addressed, when possible, by additionalprotein sources or by supplementing animal feed with non-cereal protein or synthetic amino acids. A number of strategies have been suggested to make cereal flours more complete and balanced sources of amino acids, although systematic examination of such strategies is rare. This project proposed to begin such a systematic examination using four complementary and parallel approaches to increasing wheat seed lysine: 1) Modifying endogenous wheat seed proteins for increased lysine composition. 2) Overexpression of naturally occurring high-lysine proteins in the wheat endosperm. 3) Ectopic expression of proteins in the wheat endosperm. 4) Alteration of free lysine levels in the wheat endosperm. The results of these studies are expected to be wheat lines with increased lysine content and will establish a clearer understanding of the approaches most likely to enhance cereal seed protein quality. Progress is reported for all four objectives, with a significant foundation for further work on two of the objectives (modification of wheat storage proteins and lysine metabolism). Plans for continuing work on all four objectives are briefly outlined.
Optimization of Secondary Wastewater Reuse to Minimize Environmental Risks
Authors: Oron Gideon , Mandelbaum Raphi , Enriquez Carlos E. , Armon Robert , Manor Yoseph , Gillerman L. , Alum A. , Gerba Charles P.
Published: December 2021

The main purpose of the research was to examine approaches and to evaluate methods for minimizing the risks during applying treated domestic wastewater for agricultural irrigation. This general purpose consisted of examining under field conditions the possibilities when implementing different application technologies for minimizing health and environmental risks. It was assumed that Subsurface Drip Irrigation (SDI) will provide adequate conditions for safe effluent reuse. Controlled field experiments where conducted in commercial fields to evaluate the alternatives. Main efforts where conducted in Israel in the grape vineyard in Arad heights, in the field crops in Kibbutz Chafets Chaim and in Arizona in fields adjacent to the University campus. The complementary part was to examine the behavior of the various pathogens in the effluent-soil-plant system. The analysis is based on controlled experiments, primarily in greenhouse along with field experiments. Molecular biology methods were used to identify the behavior of the pathogens in the components of the system. The project included as well examining the effluent quality in various sites, primarily those in which treated wastewater is reused for agricultural irrigation. The monitoring included conventional parameters however, also parasites such as Giardia and Cryptosporidium. The results obtained indicate the prominent advantages of using Subsurface Drip Irrigation (SDI) method for minimizing health and environmental risks during application of secondary effluent. A theoretical model for assessing the risks while applying treated wastewater was completed as well. The management model shows the risks during various scenarios of wastewater quality, application technology and related human exposure.
Interploid Hybridization of Cucumis melo and C. metuliferus
Authors: Adelberg Jeff , Skorupska Halina , Rhodes Bill , Cohen Yigal , Perl-Treves Rafael
Published: December 2021

The long-term motivation for this research is to transfer useful traits from a broad based gene pool of wild species into the narrow base of a cultivated crop in Cucumis. Our primary focus was to use polyploid prior to fertilization as a tool to overcome fertility barriers in the cross between C. melo and C. metuliferus. In conducting this research, we explored all combinations of tetraploid and diploid parents, in reciprocal combinations. Pollinations were made in both the field and greenhouse, using emasculated flowers, moneocious females, and open pollination by insect vectors, with morphological selection criteria. After observations of thousands of ovaries, we still have no definitive proof that this hybridization yielded viable embryos. The most promising results came from using tetraploid C. metuliferus, as the maternal parent in the interspecific hybridization, that set fruit were seeds contained small embryos that did not germinate. To obtain fruit set, it was important to rear plants in a cooler sunny greenhouse, as would be found in late winter/early spring. A second interspecific hybrid between wild and cultivated Cucumis, C. hystrix x C. sativus, yielded fertile progeny for the first time, while concomitantly working toward our primary goal. Two distinct treatments were necessary; 1) special plant husbandry was necessary to have the wild species produce fruit in cultivation, and 2) embryo rescue followed by chromosome doubling in vitro was required for fertility restoration. Backcrosses to crop species and resistance to nematodes are compelling areas for further work.
Computerized Synthesis of Information on the Scale Insects of the World
Authors: Ben-Dov Yair , Miller Douglass R. , Gibson G. , Kosztarab M. , Veilleux K.
Published: December 2021

Systematic information on all aspects of agriculture is a significant tool in finding solutions to various problems. This project was initiated to develop a searchable database on taxonomy, host plants, geographic distribution, economic importance and control of scale insects (Hemiptera: Coccoidea). A systematic database has been developed for 17 families of scale insects, namely, Aclerdidae, Asterolecaniidae, Beesoniidae, Carayonemidae, Cerococcidae, Coccidae, Conchaspididae, Dactylopiidae, Eriococcidae, Halimococcidae, Kerriidae, Lecanodiaspididae, Micrococcidae, Ortheziidae, Phenacoleachiidae, Phoenicococcidae and Pseudococcidae. These databases are now available on the Internet in ScaleNet - A Searchable Information System on Scale Insects - a Systematic Database of the Scale Insects of the World, (URL: http://www.sel.barc.usda.gov/scalenet/scalenet.htm ). Since 1997, the year in which ScaleNet was first placed on the Internet, this site became the best worldwide source of information on scale insects. The most reliable evaluation is that the site was 'visited' during 1999 by more than 50,000 users. Messages from the clientele of the site clearly indicate that it is widely used by researchers, applied entomologists, quarantine officers, students as well as the general public.
Osmotin and Osmotin-Like Proteins as a Novel Source for Phytopathogenic Fungal Resistance in Transgenic Carnation and Tomato Plants
Authors: Watad Abed A. , Hasegawa Paul Michael , Bressan Ray A. , Vainstein Alexander , Elad Yigal
Published: December 2021

The goal of this project is to enhance fungal resistance of carnation and tomato through the ectopic expression of osmotin and other pathogenesis-related (PR) proteins. The research objectives were to evaluate in vitro antifungal activity of osmotin and osmotin and other PR protein combinations against phytopathogens (including Fusarium oxysporum, Verticillium dahliae, Botrytus cinerea or Phytophthora infestans), develop protocols for efficient transformation of carnation and tomato, express PR proteins in transgenic carnation and tomato and evaluate fungal resistance of transgenic plants. Protocols for microprojectile bombardment and Agrobacterium-mediated transformation of carnation were developed that are applicable for the biotechnology of numerous commercial cultivars. Research established an efficient organogenetic regeneration system, optimized gene delivery and transgene expression and defined parameters requisite to the high frequency recovery of transgenic plants. Additionally, an efficient Agrobacterium-mediated transformation protocol was developed for tomato that is applicable for use with numerous commercial varieties. Rigorous selection and reducing the cytokinin level in medium immediately after shoot induction resulted in substantially greater frequency of adventitious shoots that developed defined stems suitable for rooting and reconstitution of transgenic plants. Transformation vectors were constructed for co-expression of genes encoding osmotin and tobacco chitinase Ia or PR-1b. Expression of osmotin, PR-1 and/or chitinase in transgenic carnation mediated a high level resistance of cv. White Sim (susceptible variety) to F. oxysporum f. sp. dianthi, race 2 in greenhouse assays. These plants are being evaluated in field tests. Comprehensive analysis (12 to 17 experiments) indicated that germination of B. cinerea conidia was unaffected by PR protein expression but germ tube elongation was reduced substantially. The disease severity was significantly attenuated by PR protein expression. Constitutive expression of osmotin in transgenic tomato increased resistance to B. cinerea, and P. infestans. Grey mold and late blight resistance was stable through the third selfed generation. The research accomplished in this project will have profound effects on the use of biotechnology to improve carnation and tomato. Transformation protocols that are applicable for efficient stable gene transfer to numerous commercial varieties of carnation and tomato are the foundation for the capacity to bioengineer these crops. The research further establishes that PR proteins provide a measure of enhanced disease resistance. However, considerations of PR protein combinations and conditional regulation and targeting are likely required to achieve; sustained level of resistance.
Manipulation of Gene Expression in the Chloroplast
Authors: Stern David , Schuster Gadi
Published: December 2021

The steady-state level of a given mRNA is determined by its rates of transcription and degradation. The stabilities of chloroplast mRNAs vary during plant development, in part regulating gene expression. Furthermore, the fitness of the organelle depends on its ability to destroy non-functional transcripts. In addition, there is a resurgent interest by the biotechnology community in chloroplast transformation due to the public concerns over pollen transmission of introduced traits or foreign proteins. Therefore, studies into basic gene expression mechanisms in the chloroplast will open the door to take advantage of these opportunities. This project was aimed at gaining mechanistic insights into mRNA processing and degradation in the chloroplast and to engineer transcripts of varying stability in Chlamydomonas reinhardtii cells. This research uncovered new and important information on chloroplast mRNA stability, processing, degradation and translation. In particular, the processing of the 3' untranslated regions of chloroplast mRNAs was shown to be important determinants in translation. The endonucleolytic site in the 3' untranslated region was characterized by site directed mutagensis. RNA polyadenylation has been characterized in the chloroplast of Chlamydomonas reinhardtii and chloroplast transformants carrying polyadenylated sequences were constructed and analyzed. Data obtained to date suggest that chloroplasts have gene regulatory mechanisms which are uniquely adapted to their post-endosymbiotic environment, including those that regulate RNA stability. An exciting point has been reached, because molecular genetic studies have defined critical RNA-protein interactions that participate in these processes. However, much remains to be learned about these multiple pathways, how they interact with each other, and how many nuclear genes are consecrated to overseeing them. Chlamydomonas is an ideal model system to extend our understanding of these areas, given its ease of manipulation and the existing knowledge base, some of which we have generated.
Increasing Mammary Protein Synthesis through Endocrine and Nutritional Signals
Authors: McGuire Mark A. , Arieli Amichai , Bruckental Israel , Bauman Dale E.
Published: December 2021

Objectives To determine endocrine factors that regulate the partitioning of amino acids by the mammary gland. To evaluate dietary flow and supply of energy and amino acids and their effects on milk protein synthesis and endocrine status. To use primary cultures of cow mammary epithelial cells to examine the role of specific factors on the rates and pattern of milk protein synthesis. Milk protein is an increasingly valuable component of milk but little is known regarding the specific hormonal and nutritional factors controlling milk protein synthesis. The research conducted for this project has determined that milk protein synthesis has the potential to be enhanced much greater than previously believed. Increases of over 25% in milk protein percent and yield were detected in studies utilizing abomasal infusion of casein and a hyperinsulinemic-euglycemic clamp. Thus, it appears that insulin, either directly or indirectly, can elicit a substantial increase in milk protein synthesis if additional amino acids are supplied. For additional amino acids, casein provided the best response even though substantial decreases in branched chain amino acids occur when the insulin clamp is utilized. Branched chain amino acids alone are incapable of supporting the enhanced milk protein output. The mammary gland can vary both blood flow and extraction efficiency of amino acids to support protein synthesis. A mammary culture system was used to demonstrate specific endocrine effects on milk protein synthesis. Insulin-like growth factor-I when substituted for insulin was able to enhance casein and a-lactalbumin mRNA. This suggests that insulin is a indirect regulator of milk protein synthesis working through the IGF system to control mammary production of casein and a-lactalbumin. Principal component analysis determined that carbohydrate had the greatest effect on milk protein yield with protein supply only having minor effects. Work in cattle determined that the site of digestion of starch did not affect milk composition alone but the degradability of starch and protein in the rumen can interact to alter milk yield. Cows fed diets with a high degree of rumen undegradability failed to specifically enhance milk protein but produced greater milk yield with similar composition. The mammary gland has an amazing ability to produce protein of great value. Research conducted here has demonstrated the unprecedented potential of the metabolic machinery in the mammary gland. Insulin, probably signaling the mammary gland through the IGF system is a key regulator that must be combined with adequate nutrition in order for maximum response.
Biotechnology of Tomato Volatiles for Flavor Improvement
Authors: Lewinsohn Efraim , Pichersky Eran , Gepstein Shimon
Published: December 2021

The main objectives of the research project were: 1. The manipulation, by genetic engineering techniques, of the terpenoid pathway in tomato fruit. Specifically, to test the hypothesis whether overexpression of linalool synthase in tomato fruits will result in the diversion of intermediates of the carotene biosynthetic pathway to linalool, demonstrating that linalool synthase is a key regulatory enzyme, and possibly improving tomato flavor. 2. The elucidation of the biochemical pathway leading to eugenol and methyl eugenol, and the manipulation of this pathway to determine key enzymes and to improve flavor in tomato. Background, conclusions and implications The different proportions of volatile components present in foods often determine their flavor properties. Two of the ten most important flavor compounds in tomatoes, linalool and eugenol, are emitted by the flowers of Clarkia breweri, (Onagraceae), a plant native to California, and are also present in sweet basil (Ocimum basilicum, Lamiaceae). We have studied the key enzymes and genes involved in the production of these flavorants. Linalool synthase, the key enzyme in linalool biosynthesis and its corresponding gene were isolated and characterized from Clarkia breweri. The gene was coupled to a fruit-specific tomato promotor (E8) and was used to transform tomatoes. The transgenic tomatoes produced S-linalool and 1-hydroxylinalool, compounds absent from the fruits of controls. The transgenesis did not adversely affect the overall appearance of the plants nor the levels of other terpenoids present such as carotenoids and vitamin E. Our work has proven that the terpenoid pathway in tomatoes can be modified by the introduction and expression of foreign genes coding for the enzymes controlling the production of monoterpenoid flavor compounds. We have also isolated novel enzymes and genes that are involved in the formation of eugenol and methyl eugenol from Clarkia breweri and basil. An EST library of basil glandular trichomes (the site of eugenol and methyl eugenol biosynthesis) was prepared. More than 1,200 genes have been preliminary characterized and a few of them have been confirmed by functional expression, to be involved in eugenol and methyl eugenol biosynthesis. These genes have augmented the still small repertoire of genes that are available to modify the aroma of agricultural produce by genetic engineering.
Development of A Real-Time, In-Situ Nitrate Sensor
Authors: Upadhyaya Shrini K. , Shaviv Abraham , Katzir Abraham , Shmulevich Itzhak , Slaughter David S.
Published: December 2021

Although nitrate fertilizers are critical for enhancing crop production, excess application of nitrate fertilizer can result in ground water contamination leading to the so called "nitrate problem". Health and environmental problems related to this "nitrate problem" have led to serious concerns in many parts of the world including the United States and Israel. These concerns have resulted in legislation limiting the amount of nitrate N in drinking water to 10mg/g. Development of a fast, reliable, nitrate sensor for in-situ application can be extremely useful in dynamic monitoring of environmentally sensitive locations and applying site-specific amounts of nitrate fertilizer in a precision farming system. The long range objective of this study is to develop a fast, reliable, real-time nitrate sensor. The specific objective of this one year feasibility study was to explore the possible use of nitrate sensor based on mid-IR spectroscopy developed at UCD along with the silver halide fiber ATR (i.e. attenuated total internal reflection) sensor developed at TAU to detect nitrate content in solution and soil paste in the presence of interfering compounds. Experiments conducted at Technion and UCD clearly demonstrate the feasibility of detecting nitrate content in solutions as well as soil pastes using mid-IR spectroscopy and an ATR technique. When interfering compounds such as carbonates, bicarbonates, organic matter etc. are present special data analysis technique such as singular value decomposition (SYD) or cross correlation was necessary to detect nitrate concentrations successfully. Experiments conducted in Israel show that silver halide ATR fiber based FEWS, particularly flat FEWS, resulted in low standard error and high coefficient of determination (i.e. R² values) indicating the potential of the flat Fiberoptic Evanescent Wave Spectroscopy (FEWS) for direct determinations of nitrate. Moreover, they found that it was possible to detect nitrate and other anion concentrations using anion exchange membranes and M1R spectroscopy. The combination of the ion-exchange membranes with fiberoptices offers one more option to direct determination of nitrate in environmental systems.
Importance of Endobiotics in Defense against Protozoan Ectoparasites of Fish
Authors: Noga Edward J. , Colorni Angelo , Levy Michael G. , Avtalion Ramy
Published: December 2021

Infectious disease is one of the most serious causes of economic loss in all sectors of aquaculture. There is a critical need to understand the molecular basis for protection against infectious disease so that safer, more reliable and more cost-effective strategies can be designed for their control. As part of this effort, the major goal of our BARD project was to determine the importance of endobiotics as a defense against protozoan ectoparasites in fish. Endobiotics, or antimicrobial polypeptides, are peptides and small proteins that are increasingly recognized as having a vital role in the innate defense of virtually all animals. One objective of our BARD project was to determine the antiparasitic potency of one specific group of endobiotics that were isolated from hybrid striped bass (Morone saxatilis x M chrysops). We found that these endobiotics, which we had previously named histone-like proteins (HLPs), exhibited potent activity against Amyloodinium and that the putative levels of HLPs in the skin were well within the levels that we found to be lethal to the parasite in vitro. We also found evidence for the presence of similar antibiotics in sea bream (Sparus aurata) and Mediterranean sea bass (Dicentrarchus labrax). We also examined the effect of chronic stress on the expression of HLP in fish and found that HLP levels were dramatically decreased after only one week of a crowding/high ammonia sublethal stress. We also began to explore the feasibility of upregulating endobiotics via immunostimulation. However, we did not pursue this objective as fully as we originally intended because we spent a much larger effort than originally anticipated on the last objective, the attempted isolation of novel endobiotics from hybrid striped bass. In this regard, we purified and identified four new peptide endobiotics. These endobiotics, which we have named piscidins (from "Pisces" meaning fish), have potent, broad-spectrum activity against a number of both fish and human pathogens. This includes not only parasites but also bacteria. We also demonstrated that these peptides are present in the mast cell. This was the first time that the mast cell, the most common tissue granulocyte in vertebrates, was shown to possess any type of endobiotic. This finding has important implications in explaining the possible function of mast cells in the immune response of vertebrates. In summary, the research we have accomplished in this BARD project has demonstrated that endobiotics in fish have potent activity against many serious pathogens in aquaculture and that there is considerable potential to use these compounds as stress indicators in aquaculture. There is also considerable potential to use some of these compounds in other areas of medicine, including treatment of serious infectious diseases of humans and animals.
functional genomic screen for new plant cytoskeletal proteins and the determination of their role in actin mediated functions and guard cells regulation
Authors: Sadot Einat , Staiger Christopher , Weizmann Zvi Kam
Published: December 2021

The original objectives of the approved proposal were: 1. To construct a YFP fused Arabidopsis cDNA library in a mammalian expression vector. 2. To infect the library into a host fibroblast cell line and to screen for new cytoskeletal associated proteins using an automated microscope. 3. Isolate the new genes. 4. Characterize their role in plants. The project was approved as a feasibility study to allow proof of concept that would entail building the YFP library and picking up a couple of positive clones using the fluorescent screen. We report here on the construction of the YFP library, the development of the automatic microscope, the establishment of the screen and the isolation of positive clones that are plant cDNAs encoding cytoskeleton associated proteins. The rational underling a screen of plant library in fibroblasts is based on the high conservation of the cytoskeleton building blocks, actin and tubulin, between the two kingdoms (80-90% homology at the level of amino acids sequence). In addition, several publications demonstrated the recognition of mammalian cytoskeleton by plant cytoskeletal binding proteins and vice versa. The major achievements described here are: 1. The development of an automated microscope equipped with fast laser auto-focusing for high magnification and a software controlling 6 dimensions; X, Y position, auto focus, time, color, and the distribution and density of the fields acquired. This system is essential for the high throughput screen. 2. The construction of an extremely competent YFP library efficiently cloned (tens of thousands of clones collected, no empty vectors detected) with all inserts oriented 5't03'. These parameters render it well representative of the whole transcriptome and efficient in "in-frame" fusion to YFP. 3. The strategy developed for the screen allowing the isolation of individual positive cDNA clones following three rounds of microscopic scans. The major conclusion accomplished from the work described here is that the concept of using mammalian host cells for fishing new plant cytoskeletal proteins is feasible and that screening system developed is complete for addressing one of the major bottlenecks of the plant cytoskeleton field: the need for high throughput identification of functionally active cytoskeletal proteins. The new identified plant cytoskeletal proteins isolated in the pilot screen and additional new proteins which will be isolated in a comprehensive screen will shed light on cytoskeletal mediated processes playing a major role in cellular activities such as cell division, morphogenesis, and functioning such as chloroplast positioning, pollen tube and root hair elongation and the movement of guard cells. Therefore, in the long run the screen described here has clear agricultural implications.
Phosphorylative Transduction of Developmental and Pathogenicity-Related Cues in Sclerotinia Sclerotiorum
Authors: Dickman Martin B. , Yarden Oded
Published: December 2021

Sclerotinia sclerotiorum (Lib.) de Bary is among the world's most successful and omnivorous fungal plant pathogens. Included in the more than 400 species of plants reported as hosts to this fungus are canola, alfalfa, soybean, sunflower, dry bean, and potato. The general inability to develop resistant germplasm with these economically important crops to this pathogen has focused attention on the need for a more detailed examination of the pathogenic determinants involved in disease development. This proposal involved experiments that examined the involvement of protein phosphorylation during morphogenesis (hyphal elongation and sclerotia formation) and pathogenesis (oxalic acid). Data obtained from our laboratories during the course of this project substantiates the fact that kinases and phosphatases are involved and important for these processes. A mechanistic understanding of the successful strategy(ies) used by S . sclerotiorum in infecting and proliferating in host plants and this linkage to fungal development will provide targets and/or novel approaches with which to design resistant crop plants including interference with fungal pathogenic development. The original objectives of this grant included: I. Clone the cyclic AMP-dependent protein kinase A (PKA) catalytic subunit gene from S.sclerotiorum and determine its role in fungal pathogenicity, OA production (OA) and/or morphogenesis (sclerotia formation). II. Clone and characterize the catalytic and regulatory subunits of the protein phosphatase PP2A holoenzyme complex and determine their role in fungal pathogenicity and/or morphogenesis as well as linkage with PKA-regulation of OA production and sclerotia formation. III. Clone and characterize the adenylate cyclase-encoding gene from S . sclerotiorum and detennine its relationship to the PKA/PP2A-regulated pathway. IV. Analyze the expression patterns of the above-mentioned genes and their products during pathogenesis and determine their linkage with infection and fungal growth.
Effects of root zone temperature, oxygen concentration, and moisture content on actual vs. potential growth of greenhouse crops
Authors: Lieth J. Heiner , Raviv Michael , Burger David W.
Published: December 2021

Soilless crop production in protected cultivation requires optimization of many environmental and plant variables. Variables of the root zone (rhizosphere) have always been difficult to characterize but have been studied extensively. In soilless production the opportunity exists to optimize these variables in relation to crop production.   The project objectives were to model the relationship between biomass production and the rhizosphere variables: temperature, dissolved oxygen concentration and water availability by characterizing potential growth and how this translates to actual growth. As part of this we sought to improve of our understanding of root growth and rhizosphere processes by generating data on the effect of rhizosphere water status, temperature and dissolved oxygen on root growth, modeling potential and actual growth and by developing and calibrating models for various physical and chemical properties in soilless production systems. In particular we sought to use calorimetry to identify potential growth of the plants in relation to these rhizosphere variables. While we did experimental work on various crops, our main model system for the mathematical modeling work was greenhouse cut-flower rose production in soil-less cultivation. In support of this, our objective was the development of a Rose crop model. Specific to this project we sought to create submodels for the rhizosphere processes, integrate these into the rose crop simulation model which we had begun developing prior to the start of this project. We also sought to verify and validate any such models and where feasible create tools that growers could be used for production management.   We made significant progress with regard to the use of microcalorimetry. At both locations (Israel and US) we demonstrated that specific growth rate for root and flower stem biomass production were sensitive to dissolved oxygen. Our work also identified that it is possible to identify optimal potential growth scenarios and that for greenhouse-grown rose the optimal root zone temperature for potential growth is around 17 C (substantially lower than is common in commercial greenhouses) while flower production growth potential was indifferent to a range as wide as 17-26C in the root zone. We had several set-backs that highlighted to us the fact that work needs to be done to identify when microcalorimetric research relates to instantaneous plant responses to the environment and when it relates to plant acclimation.   One outcome of this research has been our determination that irrigation technology in soilless production systems needs to explicitly include optimization of oxygen in the root zone. Simply structuring the root zone to be “well aerated” is not the most optimal approach, but rather a minimum level. Our future work will focus on implementing direct control over dissolved oxygen in the root zone of soilless production systems.
Gene Expression Patterns in Plants Colonized with Pathogenic and Non-pathogenic Gene Disruption Mutants of Colletotrichum
Authors: Rodriguez Russell J. , Freeman Stanley
Published: December 2021

Fungal plant pathogens are responsible for extensive annual crop and revenue losses throughout the world. To better understand why fungi cause diseases, we performed gene-disruption mutagenesis on several pathogenic Colletotrichum species and demonstrated that pathogenic isolates can be converted to symbionts expressing non-pathogenic lifestyles. One group of nonpathogenic mutants confer disease protection against pathogenic species of Col!etotrichum, Fusarium and Phytophthora; drought tolerance; and growth enhancement to host plants. These mutants have been defined as mutualists and disease resistance correlates to a decrease in the time required for hosts to activate defense systems when exposed to virulent fungi. A second group of non-pathogenic mutants did not confer disease resistance and were classified as commensals. In addition, we have demonstrated that wildtype pathogenic Colletotrichum species can express non-pathogenic lifestyles, including mutualism, on plants they colonize asymptomatically. We have been using wildtype and isogenic gene disruption mutants to characterize gene expression patterns in plants colonized with a pathogen, mutualist or commensal. The US group is contrasting genes expressed during colonization by mutuahstic and commensal mutants of C. magna and a pathogenic wildtype C. coccodes on tomato. The Israeli group is characterizing genes expressed during asymptomatic colonization of tomato by wildtype C. acutatum and a non-pathogenic mutant.To accomplish this we have been utilizing suppressive subtraction hybridization, microarray and sequencing strategies. The expected contribution of this research to agriculture in the US and Israel is: 1) understanding how pathogens colonize certain hosts asymptomatic ally will shed light on the ecology of plant pathogens which has been described as a fundamental deficiency in plant pathology; 2) identifying genes involved in symbiotically conferred disease resistance will help explain why and how pathogens cause disease, and may identify new candidate targets for developing genetically modified disease resistant crop plants.
Simultaneous Treatment of Odorants and Pathogens Emitted from Confined Animal Feeding Operations (CAFOs) by Advanced Oxidation Technologies
Authors: Koziel Jacek , Laor Yael , Zimmerman Jeffrey , Armon Robert , Hoff Steven , Ravid Uzi
Published: December 2021

A feasibility study was conducted, aiming to explore the potential effectiveness of UV/TiO2/O3 photooxidation technologies for simultaneous treatment of odorant and pathogen emissions from livestock and poultry operations. Several key parameters were tested in laboratory (US) and semi-pilot (Israel) scale conditions including: the effects of light energy dose (treatment time and light intensity), relative humidity and air temperature, UV wavelength, presence of photocatalyst (TiO2) and the presence of ozone. Removal and conversion of odor, target gases (sulfur-containing volatile organic compounds S-VOCs, volatile fatty acids (VFAs), phenolics, and ammonia), and airborne pathogens was tested. Up to 100% removal (below method detection level) of S-VOCs, VFAs, and phenolics, the overall odor, and up to 64.5% of ammonia was achieved with optimized treatment. Treatments involving deep UV band (185 nm) and photocatalyst (TiO2) were more efficient in removal/conversion of odorous gases and odor. The estimate of the operational cost of treatment was based on measured emissions of several odorous VOCs from full scale, commercial swine farm ranges from $0.15 to $0.59 per finisher pig. This figure represents significantly lower cost compared with the cost of biofiltration or air scrubbing.
Genome-Wide Association Study of Copy Number Variation and QTL for Economic Traits in Holstein Cattle
Authors: Seroussi Eyal , Liu George
Published: December 2021

Copy number variation (CNV) has been recently identified in human and other mammalian genomes and increasing awareness that CNV might be a major source for heritable variation in complex traits has emerged. Despite this, little has been published on CNVs in Holsteins. In order to fill this knowledge-gap, we proposed a genome-wide association study between quantitative trait loci (QTL) for economic traits and CNV in the Holstein cattle. The approved feasibility study was aimed at the genome-wide characterization of CNVs in Holstein cattle and at the demonstrating of their possible association with economic traits by performing the activities of preparation of DNA samples, Comparative Genomic Hybridization (CGH), initial association study between CNVs and production traits and characterization of CNVSNP associations. For both countries, 40 genomic DNA samples of bulls representing the extreme sub-populations for economically important traits were CGH analyzed using the same reference genome on a NimbleGen tiling array. We designed this array based on the latest build of the bovine genome (UMD3) with average probe spacing of 1150 bases (total number of probes was 2,166,672). Two CNV gene clusters, PLA2G2D on BTA2 and KIAA1683 on BTA7 revealed significant association with milk percentage and cow fertility, respectively, and were chosen for further characterization and verification in a larger sample using other methodologies including sequencing, tag SNPs and real time PCR (qPCR). Comparison between these four methods indicated that there is under estimation of the number of CNV loci in Holstein cattle and their complexity. The variation in sequence between different copies seemed to affect their functionality and thus the hybridization based methods were less informative than the methods that are based on sequencing. We thus conclude that large scale sequencing effort complemented by array CGH should be considered to better detect and characterize CNVs in order to effectively employ them in marker-assisted selection.
Genetic Basis of Cyprinid Herpes Virus-3 Resistance in Common Carp
Authors: David Lior , Palti Yaniv , Kotler Moshe , Hulata Gideon , Hallerman Eric M.
Published: December 2021

The goal of this project was to provide scientific and technical basis for initiating the development of breeding protocols using marker assisted selection for viral disease resistance in common carp. The specific objectives were: 1) Establishing families and characterizing the phenotypic and genetic variation of viral resistance; 2) Measuring the dynamics of immune response and developing a method to measure the long term immune memory; 3) Developing markers and generating a new genetic linkage map, which will enable initial QTL mapping; and, 4) Identifying genetic linkage of markers and candidate genes (like MHC and TLRs) with resistance to CyHV-3. The common carp is an important farmed freshwater fish species in the world. Edible carp is second only to tilapia in Israeli aquaculture production and ornamental carp (koi) is an important product in both the US and Israel. Carp industries worldwide have recently suffered enormous economic damage due to a viral disease caused by Cyprinid herpes virus 3 (CyHV-3). Aside from preventative measures, a sustainable solution to this problem will be to establish a genetic improvement program of the resistance of fish to the pathogen. The aims of the project was to take the necessary first steps towards that. The differences in survival rates after infection with CyHV-3 virus among 20 families from six types of crosses between three carp lines (two commercial lines and one wild-type carp) revealed that the wild-type carp and its crosses had a much-improved survival over the crosses of the commercial lines themselves. These crosses set the starting point for breeding of commercial strains with improved resistance. Resistant fish had lower antibody titer against the virus suggesting that resistance might depend more on the innate immunity. A set of 500 microsateliite markers was developed and the markers are currently being used for generating a genetic linkage map for carp and for identifying disease resistance QTL. Fourteen candidate immune genes, some of which were duplicated, were cloned from the carp and SNP markers were identified in them. The expression of these genes varied between tissues and suggested functional divergence of some duplicated genes. Initial association between CyHV-3 resistance and one of the genes was found when SNP alleles in these genes were tested for their segregation between susceptible and resistant progeny. The results of this project have implications to the development of viral resistant commercial carp strains and effective immunization against this aggressive disease. The genetic and immunological knowledge accumulated in this project will not only promote carp and koi production but will also contribute to a broader understanding of fish immunogenetics.
Inactivation of SGS3 as Molecular Basis for RNA Silencing Suppression by TYLCV V2
Authors: Gafni Yedidya , Citovsky Vitaly
Published: December 2021

The Israeli isolate of Tomato yellow leaf curl geminivirus(TYLCV-Is) is a major tomato pathogen, causing extensive crop losses in Israel and in the south-eastern U.S. Yet, little is known about the molecular mechanisms of its interaction with tomato cells. One of the most interesting aspects of such interaction is how the invading virus counteracts the RNA silencing response of the plant. In the former BARD project, we have shown that TYLCV-Is V2 protein is an RNA silencing suppressor, and that this suppression is carried out via the interaction of V2 with the SGS3 component of the plant RNA silencing machinery. This reported project was meant to use our data as a foundation to elucidate the molecular mechanism by which V2 affects the SGS3 activity. While this research is likely to have an important impact on our understanding of basic biology of virus-plant interactions and suppression of plant immunity, it also will have practical implications, helping to conceive novel strategies for crop resistance to TYLCV-Is. Our preliminary data in regard to V2 activities and our present knowledge of the SGS3 function suggest likely mechanisms for the inhibitory effect of V2 on SGS3. We have shown that V2 possess structural and functional hallmarks of an F-box protein, suggesting that it may target SGS3 for proteasomal degradation. SGS3 contains an RNA-binding domain and likely functions to protect the cleavage produces of the primary transcript for subsequent conversion to double-stranded forms; thus, V2 may simply block the RNA binding activity of SGS3. V2 may also employ a combination of these mechanisms. These and other possibilities were tested in this reported project.
Role of the viral AlkB homologs in RNA repair
Authors: Mawassi Munir , Dolja Valerian V.
Published: December 2021

AlkB proteins that repair DNA via reversing methylation damage are conserved in a broad range of prokaryotes and eukaryotes including plants. Surprisingly, AlkB-domains were discovered in the genomes of numerous plant positive-strand RNA viruses, majority of which belong to the family Flexiviridae. The major goal of this research was to reveal the AlkB functions in the viral infection cycle using a range of complementary genetic and biochemical approaches. Our hypotheses was that AlkB is required for efficient replication and genetic stability of viral RNA genomes The major objectives of the research were to identify the functions of GVA AlkB domain throughout the virus infection cycle in N. benthamiana and grapevine, to investigate possible RNA silencing suppression activity of the viral AlkBs, and to characterize the RNA demethylation activity of the mutated GVA AlkBs in vitro and in vivo to determine methylation status of the viral RNA. Over the duration of project, we have made a very substantial progress with the first two objectives. Because of the extreme low titer of the virus particles in plants infected with the AlkB mutant viruses, we were unable to analyze RNA demethylation activity and therefore had to abandon third objective. The major achievements with our objectives were demonstration of the AlkB function in virus spread and accumulation in both experimental and natural hosts of GVA, discovery of the functional cooperation and physical interaction between AlkB and p10 AlkB in suppression of plant RNA silencing response, developing a powerful virus vector technology for grapevine using GLRaV-2-derived vectors for functional genomics and pathogen control in grapevine, and in addition we used massive parallel sequencing of siRNAs to conduct comparative analysis of the siRNA populations in grape plants infected with AlkB-containing GLRaV-3 versus GLRaV-2 that does not encode AlkB. This analysis revealed dramatically reduced levels of virus-specific siRNAs in plants infected with GLRaV-3 compared to that in GLRaV-2 infection implicating AlkB in suppression of siRNA formation. We are pleased to report that BARD funding resulted in 5 publications directly supported by BARD, one US patent, and 9 more publications also relevant to project. Moreover, two joint manuscripts that summarize work on GVA AlkB (led by Israeli PI) and on viral siRNAs in grapevine (led by US PI in collaboration with University of Basel) are in preparation.