Seed coating with phages for sustainable plant biocontrol of plant pathogens and influence of the seed coat mucilage.

Pathogens resistant to classical control strategies pose a significant threat to crop yield, with seeds being a major transmission route. Bacteriophages, viruses targeting bacteria, offer an environmentally sustainable biocontrol solution. In this study, we isolated and characterized two novel phages, Athelas and Alfirin, which infect Pseudomonas syringae and Agrobacterium fabrum, respectively, and included the recently published Pfeifenkraut phage infecting Xanthomonas translucens. Using a simple immersion method, phages coated onto seeds successfully lysed bacteria post air-drying. The seed coat mucilage (SCM), a polysaccharide-polymer matrix exuded by seeds, plays a critical role in phage binding. Seeds with removed mucilage formed five to 10 times less lysis zones compared to those with mucilage. The podovirus Athelas showed the highest mucilage dependency. Phages from the Autographiviridae family also depended on mucilage for seed adhesion. Comparative analysis of Arabidopsis SCM mutants suggested the diffusible cellulose as a key component for phage binding. Long-term activity tests demonstrated high phage stability on seed surfaces and significantly increasing seedling survival rates in the presence of pathogens. Using non-virulent host strains enhanced phage presence on seeds but also has potential limitations. These findings highlight phage-based interventions as promising, sustainable strategies for combating pathogen resistance and improving crop yield.

Reduction in vertical transmission rate of bean common mosaic virus in bee-pollinated common bean plants.

Vertical transmission, the transfer of pathogens across generations, is a critical mechanism for the persistence of plant viruses. The transmission mechanisms are diverse, involving direct invasion through the suspensor and virus entry into developing gametes before achieving symplastic isolation. Despite the progress in understanding vertical virus transmission, the environmental factors influencing this process remain largely unexplored. We investigated the complex interplay between vertical transmission of plant viruses and pollination dynamics, focusing on common bean (Phaseolus vulgaris). The intricate relationship between plants and pollinators, especially bees, is essential for global ecosystems and crop productivity. We explored the impact of virus infection on seed transmission rates, with a particular emphasis on bean common mosaic virus (BCMV), bean common mosaic necrosis virus (BCMNV), and cucumber mosaic virus (CMV). Under controlled growth conditions, BCMNV exhibited the highest seed transmission rate, followed by BCMV and CMV. Notably, in the field, bee-pollinated BCMV-infected plants showed a reduced transmission rate compared to self-pollinated plants. This highlights the influence of pollinators on virus transmission dynamics. The findings demonstrate the virus-specific nature of seed transmission and underscore the importance of considering environmental factors, such as pollination, in understanding and managing plant virus spread.

Pathogenicity analysis and seed transmission of watermelon virus A in bottle gourd.

Seed transmission is among the primary strategies utilized by plant viruses for long-distance dissemination, leading to the widespread occurrence of viral diseases globally. Watermelon virus A (WVA) is a novel wamavirus first found in watermelon. However, the pathogenicity and transmission mode of WVA are still unclear. Our previous work found that the incidence of WVA in bottle gourd is very high. Based on that, the pathogenicity and seed transmission mode of WVA in bottle gourd were studied. Compared with healthy plant, bottle gourd infected by WVA showed no visible disease symptom. Moreover, in the seeds of 20 bottle gourd cultivars, the occurrence of WVA varies from 0 to 90%, and one cultivar even reaches 100%. We also found that the transmission rate from seeds to the resulting seedlings was 100%. Furthermore, WVA was present in both the seed coat and embryo, and seed disinfection cannot eliminate WVA. Besides the seed and leaf, WVA can also be detected in stem, flower, and fruit, but not in the root. To our surprise, the level of transmission from WVA-infected plants to seeds was more than 85%. In addition, the viral accumulations of both WVA and CGMMV were increased in plants with co-infection of WVA and CGMMV. Taken together, these findings reveal that WVA is a seed-transmitted virus which causes no disease symptom in bottle gourd, and there may be synergism between WVA and CGMMV.

Molecular characterization and host reaction to tomato mottle mosaic virus isolated from sweet pepper seeds in Japan.

Many countries have identified tomato mottle mosaic virus (ToMMV) as a serious threat to tomato production. Here, we constructed and characterized infectious clones of ToMMV isolated from Japanese sweet pepper seeds. The genome of the Japanese isolate is 6399 nucleotides in length and exhibits the highest identity with previously characterized isolates. For example, it is 99.7% identical to that of the Mauritius isolate, which occurs worldwide. Phylogenetic analysis based on complete genome sequences revealed that the Japanese isolates clustered in the same clade as those from other countries. When homozygous tomato cultivars with tobamovirus resistance genes were inoculated with an infectious cDNA clone of ToMMV, the virus systemically infected tomato plants with symptoms typical of Tm-1-carrying tomato cultivars. In contrast, tomato cultivars carrying Tm-2 or Tm-22 showed symptoms only on the inoculated leaves. Furthermore, when commercial cultivars of Tm-22 heterozygous tomato were inoculated with ToMMV, systemic infections were observed in all cultivars, with infection frequencies ranging from 25 to 100%. Inoculation of heterozygous sweet pepper cultivars with tobamovirus resistance genes (L1, L3, and L4) with ToMMV resulted in an infection frequency of about 70%, but most of the infected L1, L3, and L4 cultivars were symptomless, and 10-20% showed symptoms of necrosis and yellowing. Tomato mosaic virus strain L11A, an attenuated virus, did not provide cross-protection against ToMMV and led to systemic infection with typical symptoms. These results suggest that ToMMV might cause extensive damage to existing tomato and sweet pepper cultivars commonly grown in Japan.

Transmission, localization, and infectivity of seedborne maize chlorotic mottle virus.

Maize lethal necrosis is a destructive virus disease of maize caused by maize chlorotic mottle virus (MCMV) in combination with a virus in the family Potyviridae. Emergence of MLN is typically associated with the introduction of MCMV or its vectors and understanding its spread through seed is critical for disease management. Previous studies suggest that although MCMV is detected on seed, the seed transmission rate of this virus is low. However, mechanisms influencing its transmission are poorly understood. Elucidating these mechanisms is crucial for informing strategies to prevent spread on contaminated seed. In this study, we evaluated the rate of MCMV seed transmission using seed collected from plants that were artificially inoculated with MCMV isolates from Hawaii and Kenya. Grow-out tests indicated that MCMV transmission through seed was rare, with a rate of 0.004% among the more than 85,000 seed evaluated, despite detection of MCMV at high levels in the seed lots. To understand factors that limit transmission from seed, MCMV distribution in seed tissues was examined using serology and immunolocalization. The virus was present at high levels in maternal tissues, the pericarp and pedicel, but absent from filial endosperm and embryo seed tissues. The ability to transmit MCMV from seed to uninfected plants was tested to evaluate virus viability. Transmission was negatively associated with both seed maturity and moisture content. Transmission of MCMV from infested seed dried to less than 15% moisture was not detected, suggesting proper handling could be important for minimizing spread of MCMV through seed.

Localization and Mechanical Transmission of Tomato Brown Rugose Fruit Virus in Tomato Seeds.

Tomato brown rugose fruit virus (ToBRFV), belonging to the genus Tobamovirus, is a highly virulent emerging virus, causing disease outbreaks and significant crop losses worldwide. The growing number of ToBRFV epidemic episodes prompted the investigation of the role of seeds in the dissemination of the virus as an important aspect in the overall disease management. Therefore, the objectives of this study were to determine the localization of ToBRFV within tomato seeds and to evaluate its seed transmission characteristics. Seeds extracted from naturally ToBRFV-infected tomato fruits were tested for the presence of the virus using serological, molecular, and biological assays. Three immunolocalization techniques were used to determine the localization and distribution of ToBRFV within the different tissues and parts of tomato seeds. To evaluate seed transmission of ToBRFV, two grow-out experiments were conducted to assess the rate of both vertical (seeds to progeny seedlings) and possible horizontal transmission (plant to plant) based on serological and molecular assays. Seeds extracted from ToBRFV-infected fruits had a 100% contamination rate. The localization of ToBRFV in tomato seeds is only external on the seed coat (testa). Seed transmission rate from seeds to their seedlings was very low (0.08%), while no transmission was recorded from plants to plants in a small-scale greenhouse experimental setup. In conclusion, ToBRFV is a seedborne virus located externally on tomato seed coat and transmitted mechanically from ToBRFV-contaminated tomato seeds to seedlings, which could initiate a disease foci and eventually drive further dissemination and spread of the disease in a new growing area.

Kinship networks of seed exchange shape spatial patterns of plant virus diversity.

By structuring farmers' informal networks of seed exchange, kinship systems play a key role in the dynamics of crop genetic diversity in smallholder farming systems. However, because many crop diseases are propagated through infected germplasm, local seed systems can also facilitate the dissemination of seedborne pathogens. Here, we investigate how the interplay of kinship systems and local networks of germplasm exchange influences the metapopulation dynamics of viruses responsible for the cassava mosaic disease (CMD), a major threat to food security in Africa. Combining anthropological, genetic and plant epidemiological data, we analyzed the genetic structure of local populations of the African cassava mosaic virus (ACMV), one of the main causal agents of CMD. Results reveal contrasted patterns of viral diversity in patrilineal and matrilineal communities, consistent with local modes of seed exchange. Our results demonstrate that plant virus ecosystems have also a cultural component and that social factors that shape regional seed exchange networks influence the genetic structure of plant virus populations.

Under siege: virus control in plant meristems and progeny.

In the arms race between plants and viruses, two frontiers have been utilized for decades to combat viral infections in agriculture. First, many pathogenic viruses are excluded from plant meristems, which allows the regeneration of virus-free plant material by tissue culture. Second, vertical transmission of viruses to the host progeny is often inefficient, thereby reducing the danger of viral transmission through seeds. Numerous reports point to the existence of tightly linked meristematic and transgenerational antiviral barriers that remain poorly understood. In this review, we summarize the current understanding of the molecular mechanisms that exclude viruses from plant stem cells and progeny. We also discuss the evidence connecting viral invasion of meristematic cells and the ability of plants to recover from acute infections. Research spanning decades performed on a variety of virus/host combinations has made clear that, beside morphological barriers, RNA interference (RNAi) plays a crucial role in preventing-or allowing-meristem invasion and vertical transmission. How a virus interacts with plant RNAi pathways in the meristem has profound effects on its symptomatology, persistence, replication rates, and, ultimately, entry into the host progeny.

Optimal Control of Plant Disease Epidemics with Clean Seed Usage.

The distribution and use of pathogen-free planting material ("clean seeds") is a promising method to control plant diseases in developing countries. We address the question of minimizing disease prevalence in plants through the optimal usage of clean seeds. We consider the simplest possible S-I model together with a simple economic criterion to be maximized. The static optimization problem shows a diversity of possible outcomes depending on economical and epidemiological parameters. We derive a simple condition showing to what extent subsidizing clean seeds relative to the epidemiological features of the disease may help eradicate or control the disease. Then we consider dynamic optimal control and Pontryagin's maximum principle to study the optimal usage of clean seeds to control the disease. The dynamical results are comparable to the static ones and are even simpler in some sense. In particular, the condition on the critical subsidy rate that makes clean seed usage economically viable is unchanged from the static optimization case. We discuss how these results may apply to the control of maize lethal necrosis in East-Africa.

Electron Beam Susceptibility of Enteric Viruses and Surrogate Organisms on Fruit, Seed and Spice Matrices.

The objective of this study was to use high-energy electron beam (HEEB) treatments to find surrogate microorganisms for enteric viruses and to use the selected surrogates as proof of concept to investigate low-energy electron beam (LEEB) treatments for enteric virus inactivation at industrial scale on frozen blueberries. Six food matrices inoculated with HAV (hepatitis A virus), MNV S99 (murine norovirus), bacteriophages MS2 and Qß, and Geobacillus stearothermophilus spores were treated with HEEB at 10 MeV using 4, 8 and 16 kGy doses. G. stearothermophilus spores showed the highest inactivation on all matrices except on raisins, with a dose-dependent effect. HAV reached the maximum measurable log10 reduction (> 3.2 log10) when treated at 16 kGy on raisins. MNV showed the highest resistance of all tested microorganisms, independent of the dose, except on frozen blueberries. On frozen blueberries, freeze-dried raspberries, sesame seeds and black peppercorns, HAV showed a mean inactivation level in between those of MS2 and G. stearothermophilus. Based on this, we selected both surrogate organisms as first approximation to estimate HAV inactivation on frozen blueberries during LEEB treatment at 250 keV using 16 kGy. Reductions of 3.1 and 1.3 log10 were measured for G. stearothermophilus spores and MS2, respectively, suggesting that a minimum reduction of 1.4 log10 can be expected for HAV under the same conditions.

Development and validation of a real-time RT-PCR test for screening pepper and tomato seed lots for the presence of pospiviroids.

Potato spindle tuber viroid and other pospiviroids can cause serious diseases in potato and tomato crops. Consequently, pospiviroids are regulated in several countries. Since seed transmission is considered as a pathway for the introduction and spread of pospiviroids, some countries demand for the testing of seed lots of solanaceous crops for the presence of pospiviroids. A real-time RT-PCR test, named PospiSense, was developed for testing pepper (Capsicum annuum) and tomato (Solanum lycopersicum) seeds for seven pospiviroid species known to occur naturally in these crops. The test consists of two multiplex reactions running in parallel, PospiSense 1 and PospiSense 2, that target Citrus exocortis viroid (CEVd), Columnea latent viroid (CLVd), pepper chat fruit viroid (PCFVd), potato spindle tuber viroid (PSTVd), tomato apical stunt viroid (TASVd), tomato chlorotic dwarf viroid (TCDVd) and tomato planta macho viroid (TPMVd, including the former Mexican papita viroid). Dahlia latent viroid (DLVd) is used as an internal isolation control. Validation of the test showed that for both pepper and tomato seeds the current requirements of a routine screening test are fulfilled, i.e. the ability to detect one infested seed in a sample of c.1000 seeds for each of these seven pospiviroids. Additionally, the PospiSense test performed well in an inter-laboratory comparison, which included two routine seed-testing laboratories, and as such provides a relatively easy alternative to the currently used tests.

Maize lethal necrosis (MLN): Efforts toward containing the spread and impact of a devastating transboundary disease in sub-Saharan Africa.

Maize lethal necrosis (MLN), a complex viral disease, emerged as a serious threat to maize production and the livelihoods of smallholders in eastern Africa since 2011, primarily due to the introduction of maize chlorotic mottle virus (MCMV). The International Maize and Wheat Improvement Center (CIMMYT), in close partnership with national and international partners, implemented a multi-disciplinary and multi-institutional strategy to curb the spread of MLN in sub-Saharan Africa, and mitigate the impact of the disease. The strategy revolved around a) intensive germplasm screening and fast-tracked development and deployment of MLN-tolerant/resistant maize hybrids in Africa-adapted genetic backgrounds; b) optimizing the diagnostic protocols for MLN-causing viruses, especially MCMV, and capacity building of relevant public and private sector institutions on MLN diagnostics and management; c) MLN monitoring and surveillance across sub-Saharan Africa in collaboration with national plant protection organizations (NPPOs); d) partnership with the private seed sector for production and exchange of MLN pathogen-free commercial maize seed; and e) awareness creation among relevant stakeholders about MLN management, including engagement with policy makers. The review concludes by highlighting the need to keep continuous vigil against MLN-causing viruses, and preventing any further spread of the disease to the major maize-growing countries that have not yet reported MLN in sub-Saharan Africa.

The threat of seed-transmissible pepper yellow leaf curl Indonesia virus in chili pepper.

Recently, chili pepper (Capsicum annuum) plants in Indonesia have been devastated by a notorious bipartite begomovirus infection named Pepper yellow leaf curl Indonesia virus (PepYLCIV), which causes a distinct decrease in chili pepper production. Pepper yellow diseases have been known since early 2000; however, the spread of this virus thus far is distressing. These diseases can reduce chili yields by 20-100% in Indonesia. As previously known, begomovirus can be transmitted through whitefly to several host plants from the families Solanaceae, Compositae, and Leguminosae. In the field, a single plant was observed with severe symptoms of pepper yellow leaf curl disease, while other plants in the same field were asymptomatic and healthy. The observation leads to the possibility that the virus can be transmitted from previously infected chili pepper plants through seeds, as begomovirus transmission through seeds has been reported before. This study was conducted using seeds from chili peppers infected with viruses from different places in Indonesia. Whole seeds, embryos, and seedlings from PepYLCIV infected seeds were investigated in this study by performing viral genome DNA extraction, uracil DNA glycosylase-PCR, and sequencing analysis. Results revealed that both DNA-A and DNA-B of PepYLCIV in seeds and embryos of infected chili pepper plants were detected. The results also showed that 25-67% of PepYLCIV DNA-A and 50-100% of DNA-B were detected from seedlings grown from infected chili pepper seed collected from different location, thus confirming PepYLCIV as a seed-transmissible virus in chili pepper plants.

Seed transmission of a distinct soybean yellow mottle mosaic virus strain identified from India in natural and experimental hosts.

Soybean yellow mottle mosaic virus (SYMMV) is a newly identified member of the genus Gammacarmovirus from grain legumes in India. As the modes of transmission of this virus have not been described, we assessed the possibility of SYMMV to be transmitted through seed collected from field infected mungbean plants and mechanically sap inoculated French bean plants using serological and molecular techniques followed by progeny assays. Direct antigen coated enzyme linked immunosorbent assay (DAC-ELISA) and reverse transcription polymerase chain reaction (RT-PCR) results are inconsistent with field infected mungbean seed tissues to ensure seed transmissibility irrespective of seed number used. Seed from mechanical sap inoculated French bean showed higher absorbance values in DAC-ELISA and amplification corresponding to replicase, movement and coat protein regions of SYMMV genome. The relative accumulation of SYMMV was higher in pod walls, immature seed and stamens and stigma of mechanical sap inoculated French bean. Progeny assays with infected seed revealed the seed transmissibility of SYMMV at the rate of 63.33% in mungbeanand 73.33% in French bean. Mechanical sap inoculation of mungbean progeny seedlings on French bean cv. Pusa Parvati produced characteristic symptoms of SYMMV. The results obtained from this study demonstrate that SYMMV is seed borne in nature and can be transmitted to next generation seedlings. This is the first report of seed transmission of SYMMV in mungbean and French bean.

Impact of Turnip yellows virus infection on seed yield of an open-pollinated and hybrid canola cultivar when inoculated at different growth stages.

Turnip yellows virus (TuYV; family Luteoviridae, genus Polerovirus) is the most economically damaging virus infecting canola (Brassica napus) in the south-west Australian grainbelt. However, the impact of TuYV infection at different growth stages on canola seed yield has not been examined. This information is vital for implementing targeted management strategies. Four glasshouse experiments were conducted to examine seed yield losses incurred by an open-pollinated (ATR Bonito) and hybrid (Hyola® 404RR) canola cultivar when aphid-inoculated with TuYV at GS12 (two leaves unfolded), GS17 (seven leaves unfolded), GS30 (beginning of stem elongation) and GS65 (full flowering). When inoculated at GS12 and GS17, cv. Bonito plants incurred 30 % and 36 % seed yield losses, respectively, compared to healthy plants. Similarly, cv. 404RR incurred 41 % and 26 % seed yield losses at GS12 and GS17, respectively. However, when inoculated at GS30, whilst cv. Bonito plants incurred a 26 % seed yield loss, cv. 404RR incurred no significant loss. Neither cultivar incurred seed yield losses from inoculation at GS65. Additional information was collected from these experiments to improve sampling protocols to enhance TuYV detection, with a molecular and serological technique. When canola plants were at pre-flowering growth stages, TuYV was reliably detected 7-14 days after inoculation (DAI) in the youngest leaf. Once flowering had begun, TuYV was consistently detected 7-14 DAI in petals and flower buds. In contrast, regardless of growth stage, testing the oldest leaf regularly resulted in delayed detection or false negatives. Information generated in this study helps to quantify the value of management strategies targeted at preventing TuYV spread in pre-flowering canola crops and ultimately increase the efficiency of resource use.

Potential use of newly isolated bacteriophage as a biocontrol against Acidovorax citrulli.

Acidovorax citrulli, the gram-negative bacteria that causes bacterial fruit blotch (BFB), has been responsible for huge worldwide economic losses in watermelon and melon production since 1980. No commercial cultivar resistant to BFB has been reported. Of the two reported genotypes of A. citrulli, genotype I is the main causal agent of BFB in melon and genotype II causes disease in watermelon. After the isolation of the first bacteriophage against A. citrulli (ACP17), efforts have been made to isolate bacteriophages with wider host ranges by collecting samples from watermelon, pumpkin, and cucumber. The newly isolated phage ACPWH, belonging to the Siphoviridae family, has a head size of 60 ± 5 nm and tail size of 180 ± 5 nm, and can infect 39 out of 42 A. citrulli strains. ACPWH has genome size of 42,499 and GC content of 64.44%. Coating watermelon seeds with bacteriophage ACPWH before soil inoculation with A. citrulli resulted in 96% germination and survival, compared to 13% germination of uncoated control seeds. These results suggest that phage ACPWH may be an effective and low-cost biocontrol agent against BFB.

Quantitative Real-Time PCR Analysis of Individual Flue-Cured Tobacco Seeds and Seedlings Reveals Seed Transmission of Tobacco Mosaic Virus.

Tobacco mosaic virus (TMV) is an extensively studied RNA virus known to infect tobacco (Nicotiana tabacum) and other solanaceous crops. TMV has been classified as a seedborne virus in tobacco, with infection of developing seedlings thought to occur from contact with the TMV-infected seed coat. The mechanism of TMV transmission through seed was studied in seed of the K 326 cultivar of flue-cured tobacco. Cross pollinations were performed to determine the effect of parental tissue on TMV infection in seed. Dissection of individual tobacco seeds into seed coat, endosperm, and embryo was performed to determine TMV location within a seed, while germination tests and separation of the developing seedling into seed coat, roots, and cotyledons were conducted to estimate the percent transmission of TMV. A reverse-transcriptase quantitative PCR (RT-qPCR) assay was developed and used to determine TMV concentrations in individual seed harvested from pods that formed on plants from TMV-infected and noninfected crosses. The results showed maternal transmission of TMV to tobacco seed and seedlings that developed from infected seed, not paternal transmission. RT-qPCR and endpoint PCR assays were also conducted on the separated seed coat, endosperm, and embryo of individual seed and separated cotyledons, roots, and seed coats of individual seedlings that developed from infected tobacco seed to identify the location of the virus in the seed and the subsequent path the virus takes to infect the developing seedling. RT-qPCR and endpoint PCR assay results showed evidence of TMV infection in the endosperm and embryo, as well as in the developing seedling roots and cotyledons within 10 days of initiating seed germination. To our knowledge, this is the first report of TMV being detected in embryos of tobacco seed, demonstrating that TMV is seedborne and seed-transmitted in flue-cured tobacco.

Revisiting Seed Transmission of the Type Strain of Tomato yellow leaf curl virus in Tomato Plants.

Isolates of the Tomato yellow leaf curl virus (TYLCV) species (genus Begomovirus, family Geminiviridae) infect tomato crops worldwide, causing severe economic damage. Members of the whitefly Bemisia tabaci sibling species group are the vector of begomoviruses, including TYLCV. However, transmission of isolates of the type strain (Israel [IL]) of TYLCV (TYLCV-IL) by tomato seed has recently been reported based on infections occurring in Korea. Because of the consequences of this finding on the epidemiology and control of the disease caused by TYLCV and on the seed market, it was considered essential to revisit and expand those results to other tomato-growing areas. TYLCV DNA content was detected in tomato and Nicotiana benthamiana seed collected from plants naturally or experimentally infected with TYLCV-IL, supporting its seedborne nature. The TYLCV-IL replication detected in tomato and N. benthamiana flower reproductive organs demonstrated close association of this virus with the seed during maturation. However, the significant reduction of TYLCV DNA load after surface disinfections of tomato seed suggests that most of the virus is located externally, as contaminant of the seed coat. Transmission assays, carried out with seven tomato genotypes and more than 3,000 tomato plants, revealed no evidence of seed transmission from "surface-disinfected" or untreated seed for two Mediterranean isolates of TYLCV-IL. Similar results were also obtained for seed collected from TYLCV-IL-infected N. benthamiana plants. The results support the conclusion that TYLCV-IL is seedborne but is not seed transmitted in tomato or N. benthamiana, suggesting that transmission through seed is not a general property of TYLCV.

Genome-Wide Analyses and Prediction of Resistance to MLN in Large Tropical Maize Germplasm.

Maize lethal necrosis (MLN), caused by co-infection of maize chlorotic mottle virus and sugarcane mosaic virus, can lead up to 100% yield loss. Identification and validation of genomic regions can facilitate marker assisted breeding for resistance to MLN. Our objectives were to identify marker-trait associations using genome wide association study and assess the potential of genomic prediction for MLN resistance in a large panel of diverse maize lines. A set of 1400 diverse maize tropical inbred lines were evaluated for their response to MLN under artificial inoculation by measuring disease severity or incidence and area under disease progress curve (AUDPC). All lines were genotyped with genotyping by sequencing (GBS) SNPs. The phenotypic variation was significant for all traits and the heritability estimates were moderate to high. GWAS revealed 32 significantly associated SNPs for MLN resistance (at p < 1.0 × 10-6). For disease severity, these significantly associated SNPs individually explained 3-5% of the total phenotypic variance, whereas for AUDPC they explained 3-12% of the total proportion of phenotypic variance. Most of significant SNPs were consistent with the previous studies and assists to validate and fine map the big quantitative trait locus (QTL) regions into few markers' specific regions. A set of putative candidate genes associated with the significant markers were identified and their functions revealed to be directly or indirectly involved in plant defense responses. Genomic prediction revealed reasonable prediction accuracies. The prediction accuracies significantly increased with increasing marker densities and training population size. These results support that MLN is a complex trait controlled by few major and many minor effect genes.

Prevalences of Pospiviroid Contamination in Large Seed Lots of Tomato and Capsicum, and Related Seed Testing Considerations.

Analyses of pospiviroids in commercial seed lots of tomato and capsicum, determined by testing of 12,000 to 40,000 seeds per lot, have enabled the development of empirically-derived distribution curves for the observed prevalences of viroids in those commodities. Those distribution curves can be considered in conjunction with statistically-based estimates of detection that would be achieved using other sample sizes. Statistical calculations using binomial distributions show that sample sizes of 3000 and 9400 seeds allow detection of viroid prevalences as low as 0.1% and 0.032%, respectively, with 95% confidence. Applying those calculations to observed viroid prevalences in contaminated tomato seed lots, it is estimated that the use of sample sizes of 3000 and 9400 seeds would detect 15% and 42%, respectively, of the contaminated seed lots identified using the larger sample sizes of approximately 20,000 seeds reported in this study. It is concluded that the higher costs associated with testing of larger sample sizes represent a worthwhile investment in agricultural biosecurity.