Sue A. Tolin

Genetic characteristics of two genes for resistance to soybean mosaic virus in PI486355 soybean

Soybean [Glycine max (L.) Merr.] PI486355 is resistant to all the identified strains of soybean mosaic virus (SMV) and possesses two independently inherited resistance genes. To characterize the two genes, PI486355 was crossed with the susceptible cultivars ‘Lee 68’ and ‘Essex’ and with cultivars ‘Ogden’ and ‘Marshall’, which are resistant to SMV-G1 but systemically necrotic to SMV-G7. The F2 populations and F2∶3 progenies from these crosses were inoculated with SMV-G7 in the greenhouse. The two resistance genes were separated in two F3∶4 lines, ‘LR1’ and ‘LR2’, derived from Essex x PI486355. F1 individuals from the crosses of LR1 and LR2 with Lee 68, Ogden, and ‘York’ were tested with SMV-G7 in the greenhouse; the F2 populations were tested with SMV-G1 and G7. The results revealed that expression of the gene in LR1 is gene-dosage dependent, with the homozygotes conferring resistance but the heterozygotes showing systemic necrosis to SMV-G7. This gene was shown to be an allele of the Rsv1 locus and was designated as Rsv1-s. It is the only allele identified so far at the Rsv1 locus which confers resistance to SMV-G7. Rsv1-s also confers resistance to SMV-G1 through G4, but results in systemic necrosis with SMV-G5 and G6. The gene in LR2 confers resistance to strains SMV-G1 through G7 and exhibits complete dominance. It appears to be epistatic to genes at the Rsv1 locus, inhibiting the expression of the systemic necrosis conditioned by the Rsv1 alleles. SMV-G7 induced a pin-point necrotic reaction on the inoculated primary leaves in LR1 but not in LR2. The unique genetic features of the two resistance genes from PI486355 will facilitate their proper use and identification in breeding and contribute to a better understanding of the interaction of SMV strains with soybean resistance genes.

Genetic and Phenotypic Analysis of Soybean mosaic virus Resistance in PI 88788 Soybean

ABSTRACT Resistance to Soybean mosaic virus (SMV) was identified in PI 88788 soybean, a germ plasm accession from China that is used widely as a source of resistance to soybean cyst nematode. Strains SMV-G1 through -G7 infected the inoculated leaves of PI 88788 but were not detected in upper, noninoculated trifoliolate leaves. Inheritance of resistance was determined by inoculating progenies of crosses of PI 88788 with susceptible cvs. Essex and Lee 68 with SMV strains G1 and G7. Allelomorphic relationships with known genes for resistance to SMV were tested in crosses with the resistant genotypes PI 96983, L29, and V94-5152, possessing Rsv1, Rsv3, and Rsv4 genes, respectively. Data analyses showed that resistance in PI 88788 to SMV-G1 is controlled by a single, partially dominant gene; however, to SMV-G7, the same gene was completely dominant. The PI 88788 gene was independent of the Rsv1 and Rsv3 loci, but allelic to Rsv4 in V94-5152. Expression of the Rsv4 gene in PI 88788 resulted in a reduced number of infection sites and restricted short- and long-distance movement of virus, rather than hypersensitivity. A unique late susceptible phenotype was strongly associated with heterozygosity. This gene has potential value for use in gene pyramiding to achieve resistance to several SMV strains, as well as for rate-reducing resistance.

Tissue blot immunoassay and direct RT-PCR of cucumoviruses and potyviruses from the same NitroPure nitrocellulose membrane

A method is described for using Nitropure nitrocellulose (NPN) membranes as an effective solid matrix for retrieval of template RNA of three potyviruses, Tobacco etch virus, Soybean mosaic virus and Turnip mosaic virus, and two cucumoviruses, Cucumber mosaic virus and Peanut stunt virus. These NPN membranes were also used for tissue blot immunosorbent assays (TBIAs) to identify and detect plant viruses. For RNA detection, discs from dried membranes blotted with infected tissue were minimally cleaned with Triton X-100 and placed directly into reverse transcription (RT) reactions to initiate cDNA synthesis. Aliquots of cDNA plus primers specific for coat protein produced PCR amplicons of expected sizes for each of the viruses. Intensity of PCR-amplified bands from cDNA transcribed from both non-processed and TBIA-processed NPN membranes was comparable to those using FTA Card protocols. Direct sequencing of PCR products yielded high quality runs enabling identification to species. NPN membranes retained immunologically detectable virus particles, as well as intact template viral RNA, for more than a year at room temperature. The quantity of amplification product decreased after several months of storage, but could be increased by increasing the number of PCR cycles.

Seasonal Abundance and Diversity of Aphids (Homoptera: Aphididae) in a Pepper Production Region in Jamaica

Abstract Seasonal dispersal and diversity of aphid species were monitored on pepper farms in St. Catherine, Jamaica throughout 1998 and 1999 to identify the most likely vectors of tobacco etch virus (TEV) in pepper fields. Flight activity was monitored weekly on five farms using water pan traps. More than 30 aphid species were identified, 12 of which are new records for Jamaica. Ninety-two percent of the aphids captured from October 1998 through July 1999 belonged to only seven of the >30 species identified. Of these seven species, Aphis gossypii Glover and those in the Uroleucon ambrosiae (Thomas) complex comprised more than two-thirds of the total. Five known vectors of TEV were captured: A. gossypii, Aphis craccivora Koch, Aphis spiraecola Patch, Myzus persicae (Sulzer), and Lipaphis pseudobrassicae Davis. Generally, more aphids were collected from mid-September through mid-May than from mid-May through mid-September. The influence that rainfall and temperature had on periods of aphid flight activity also was investigated. Results indicated that flight of some species increased 3–4 wk after a rainfall event, whereas temperature did not appear to affect flight activity. High populations of A. gossypii as well as the presence of four additional known TEV vectors were encountered in October and November, which is the period that significant acreage is transplanted to pepper for harvest to coincide with the winter export market. Because pepper is most vulnerable to yield loss when young plants become infected with TEV, pepper production in Jamaica may be threatened if commonly abundant species such as A. gossypii are carrying TEV. Based on this information, implications for management of pepper viruses and their aphid vectors in Jamaica are discussed.

The Status, Promise and Potential Perils of Commercially Available Genetically Modified Microorganisms in Agriculture and the Environment

Modern genetics has shown the power of modifying microbes, from viruses to bacteria to algae, to produce desirable agricultural products. Nevertheless, gene additions or modification have led to relatively few products in the marketplace due partly to costs of regulation, but also to the challenges of production, delivery and application. Some products with gene loss have been marketed, notably Agrobac-terium radiobacter with a deletion for plasmid transfer, some veterinary vaccines and plants with one or a few genes from microbes for plant protection. Concerns over using live microbes are centered on recombination with wild type strains, potential for environmental risks, market acceptance, market scope, monitoring costs, and costs of production. The challenges in microbial agricultural plant biotechnology far outweigh those in medical and veterinary biotechnology because of pricing potential, larger markets and controlled environments in which modified microbes can function. Nevertheless, the promise and need for control of plant pathogens for which little or no plant resistance is available warrant continued efforts in this area. Veterinary uses of modified microbes will continue and be more widely accepted. Plants ‘vaccinated’ with genes for plant protection are increasingly used but their safety is still questioned and debated. Products such as enzymes from GMOs will continue to enter the marketplace and be accepted with few questions.

Virus Diseases of Tropical Vegetable Crops and Their Management

Viruses have long been known to be prevalent in plants in tropical and sub-tropical developing countries, particularly in staple crops such as cassava, rice, coconut and pulses. The need to address a wider range of vegetable crops was identified by the IPM-Innovation Lab. To meet these needs, a team of virologists was organized to work across countries and regions with IPM specialists to document virus disease problems in priority crops; mainly tomato and peppers, melons and various gourds and cucurbits, locally preferred vegetables such as eggplant (brinjal), okra (bhendi) and yardlong bean, and fruits (passion fruit, tree tomato). These crops constitute important sources of income and food sources, for many farmers, and were observed to be infected by a wide diversity of viruses. Demands for increased production, increased uniformity of vegetables grown for domestic and export markets, changes in production practices leading to scale up of production of seeds and seedlings, changes related to intensification and global climate change, and greater crop uniformity across regions, appear to be associated with crop losses due to viruses. This chapter summarizes more than two decades of research results to identify problems, and describes progress to enhance local capacity for in-country diagnosis and implementation of integrated disease management practices.

Establishment and maintenance of Soybean Mosaic Virus in soybean callus culture

Susceptibility to inoculation with Soybean mosaic virus (SMV) and virus activity were investigated in soybean callus cultures growing in vitro. Excised hypocotyls of susceptible soybean, Glycine max (L.) Merr. ‘Essex’, were cultured in Msoy medium in the light at 25 °C. Established calluses were inoculated with SMV in vitro by a soak–prick method. In addition, SMV-infected leaves of soybean ‘Lee 68’ were surface sterilized, excised, and placed on callus-inducing medium. Calluses infected with SMV initiated by either method grew in vitro as well as calluses from uninfected tissues. Callus cultures turned brownish yellow after 6–8 weeks, when the media became depleted of nutrients. However, callus with an active SMV infection could be maintained by regular subculture to fresh medium. Longevity of SMV–callus cultures was increased by storage at 10–15 °C, thus reducing the frequency of transfers. The virus was detected in infected calluses by serological tests. Infectivity assays confirmed the presence and viability of SMV in callus cultures. Most callus cultures induced directly from infected leaves retained virus and high infectivity, whereas infective cultures from in vitro inoculated hypocotyls appeared to decrease in number and infectivity with repeated subcultures. However, selected infective cultures retained high infectivity after 10 successive transfers over a period of 20 months. These results demonstrate that SMV can be cultured and maintained active in callus cultures in vitro