Novel sources of turnip yellows virus resistance in Brassica and impacts of temperature on their durability

Turnip yellows virus (TuYV; family Solemoviridae, genus Polerovirus) is the most widespread and economically damaging virus of canola (Brassica napus L.) production in Australia. However, no Australian commercial seed companies currently market TuYV-resistant canola cultivars and little information is available on the susceptibility of those available. To identify potential sources of TuYV resistance, 100 B. napus accessions from the ERANET-ASSYST diversity set were screened in the field and five of these were selected for further phenotyping via aphid inoculation. Furthermore, 43 Australian canola cultivars, six B. napus genotypes with previously reported resistance, and 33 B. oleracea and B. rapa cultivars were phenotyped. All Australian cultivars were susceptible except for ATR Stingray. Stronger resistance to TuYV infection (IR) was identified in diversity set accessions Liraspa-A, SWU Chinese 3 and SWU Chinese 5. As indicated by lower relative ELISA absorbance values (R-E405) in infected plants, resistance to TuYV accumulation (AR) often accompanied IR. Moderate IR was identified in four B. oleracea and one B. rapa cultivars. Very strong AR was identified in four B. oleracea cultivars and AR of some degree was common across many cultivars of this species tested. The impact of temperature during the inoculation access period or post-inoculation incubation on the resistance identified was examined. Infection rates were significantly higher in resistant B. napus genotypes when inoculated at 16°C than at 26°C suggesting an increase in aphid transmission efficiency. IR in B. napus genotypes was strong when incubated at 16°C but weakened at elevated temperatures with almost total breakdown in most genotypes at 30°C. However, infected plants of B. napus and B. oleracea genotypes with AR maintained lower R-E405 than susceptible controls at all temperatures tested. Novel sources of resistance identified in this study offer potential as breeding material in Australia and abroad.

Transmission Efficiency of Papaya ringspot virus by Three Aphid Species

The transmission efficiency of Papaya ringspot virus (PRSV) by three aphid vectors (i.e., Aphis gossypii, A. craccivora, and Myzus persicae) was studied. Efficiency was measured by single-aphid inoculation, group inoculation (using five aphids), duration of virus retention, and the number of plants following a single acquisition access period (AAP) to which the aphids could successfully transmit the virus. Single-aphid inoculation studies indicated that M. persicae (56%) and A. gossypii (53%) were significantly more efficient in transmitting PRSV than A. craccivora (38%). Further, in the former two species, the time required for initiation of the first probe on the inoculation test plant was significantly shorter compared to A. craccivora. PRSV transmission efficiency was 100% in all three species when a group of five aphids were used per plant. There was a perceptible decline in transmission efficiency as the sequestration period increased, although M. persicae successfully transmitted PRSV after 30 min of sequestration. A simple leaf-disk assay technique was employed for evaluating the transmission efficiency of three species of aphids. The results of leaf-disk assays also indicated that A. gossypii (48%) and M. persicae (56%) were more efficient PRSV vectors than A. craccivora. Using leaf-disk assays, the ability of individual aphids to inoculate PRSV serially to a number of plants was studied. Following a single AAP on an infected leaf, M. persicae was more efficient than the other two species with 52.5% transmission after the first inoculation access period (IAP). However, its inoculation efficiency significantly decreased with the second and subsequent IAPs. A. gossypii was able to transmit PRSV sequentially up to four successive leaf disks, but with significantly declining efficiency. Since A. gossypii is reported to be the numerically dominant vector in south India in addition to being a more efficient vector capable of inoculating PRSV to multiple plants, it should be the target vector for control strategies.

Adaptation of plum pox virus to a herbaceous host (Pisum sativum) following serial passages

Plum pox virus (PPV) populations from peaches are able to adapt consistently to herbaceous hosts, characterized by a reduction in time to symptom development, increases in inoculation efficiency and increased titres. PPV adaptation was studied by using pea (Pisum sativum) as an alternative host. Two isolates of PPV from peaches were inoculated and passaged in peas ten times using either aphid or mechanical inoculation, generating four independent passage lines. Mechanical-transmission efficiency from peach to pea improved from 3 % at passage 1 to 100 % by serial passage 4 on peas. Inoculation using aphid vectors required six to ten serial passages in pea to reach a peak of 50–60 % transmission efficiency. Sequence analyses of all four PPV population lines inoculated sequentially to pea identified a specific mutation occurring consistently in the NIb gene when compared with the same PPV isolates passaged in parallel in peach. The mutation allowed PPV to replicate up to 20 times faster in the new host. Pea-adapted strains of PPV at every passage were also tested for their ability to infect the original host, peach. Regardless of the number of previous passages, all pea-adapted PPV strains consistently infected peach at low levels using aphid inoculation.

Factors Affecting the Transmission and Spread of Sugarcane yellow leaf virus

Sugarcane, Saccharum spp. hybrid, is widely infected in the United States and many other countries with a yellowing and stunting disease called sugarcane yellow leaf syndrome. The causal agent, Sugarcane yellow leaf virus (ScYLV), is a Polerovirus of the Luteoviridae family. In this study, it was transmitted by the sugarcane aphid, Melanaphis sacchari, and also by the corn leaf aphid, Rhopalosiphum maidis, and the rice root aphid, R. rufiabdominalis. Two other aphids that infest sugarcane in Hawaii did not transmit the virus. Some Hawaiian sugarcane cultivars are susceptible to ScYLV, while others remain virus-free in the field. The latter were not infected when inoculated with viruliferous M. sacchari. Virus-free plants of susceptible cultivars were produced through apical meristem culture and were readily reinfected by viruliferous M. sacchari. They were also quickly reinfected when planted in a field in proximity to other infected sugarcane naturally infested with M. sacchari. Sugarcane cultivars are hybrids of several Saccharum species. In a field-grown collection of Saccharum and related species, 11 to 71% of the clones of four of the species were infected with ScYLV. None of the related genus Erianthus plants were infected, but four clones were infected experimentally by aphid inoculation. A low to moderate percentage of corn, rice, and sorghum seedlings became infected when inoculated with ScYLV, but barley, oats, and wheat proved to be very susceptible. None of seven weeds common in sugarcane fields were infected with ScYLV.

Transfer of Plum Pox Virus Coat Protein Genes from a Plum Pox-resistant Transgenic Clone of Prunus domestica Plum to Its Progeny through Hybridization

Sharka or plum pox virus (PPV) is a major disease of stone fruit and causes severe economic losses in Europe. There is little resistance to PPV in most Prunus species, thus genetic engineering represents a potentially useful approach to obtain resistant germplasm. Transgenic plums containing the PPV coat protein (CP) or the related papaya ringspot virus (PRV)-CP gene were produced through Agrobacterium tumefaciens-mediated transformation. These transgenic plum clones were then evaluated for resistance to PPV infection in the greenhouse by graft or aphid inoculation with PPV. While symptoms of PPV appeared in most transgenic clones, all plants of PPV-CP transgenic clone C5 were symptomless and ELISA and immunocapture-reverse transcriptase PCR negative for over three years following inoculation with two strains of PPV (Ravelonandro et al., Plant Dis. 81:1231-1235, 1997). Clone C5, which contains multiple copies of the PPV-CP gene, was hybridized with PRV-CP transgenic plants or untransformed plum cultivars. Progeny were obtained containing no transgenes, only the PPV-CP, only the PRV-CP, or both the PRV-CP and PPV-CP transgenes. Seedlings were inoculated with PPV. At 5 and 11 months post-inoculation, seedlings containing the PPV-CP genes from C5 were symptomless and ELISA negative. Seedlings containing only PRV-CP transgenes or non-transformed controls showed symptoms of PPV infection and were ELISA positive. These results indicate that the PPV-CP transgenes can be transferred to progeny through hybridization and that these genes can impart resistance to PPV in transgenic seedlings. The inheritance of the multicopy inserts of the PPV-CP and PRV-CP transgenes is being analyzed. The combined effects of both transgenes on resistance to PPV and the stability of PPV resistance in the progeny of the resistant C5 transgenic line are currently under evaluation.

Resistance to Blueberry Shoestring Virus in Southern Highbush and Rabbiteye Cultivars

To determine if blueberry shoestring virus (BBSSV) is absent in the southern United States due to resistance of cultivars, we mechanically and rub-inoculated 1-year-old rooted microshoots of nine cultivars representing southern rabbiteye (Vaccinium ashei Reade), southern highbush (hybrids of V. corymbosum and V. darrowi Camp), and northern highbush (V. corymbosum L.). Leaves were sampled from plants, and enzyme-linked immunosorbent assay screened for the presence of virus over 15 months. Only a few individuals were infected after aphid inoculation, but many northern and southern cultivars became infected after mechanical inoculation. Northern highbush `Elliot' (50%) and `Blueray' (46.3%) had the highest infection rates, followed by rabbiteye `Climax' (36.3%) and the southern highbush `O'Neal' (12.5%). The lowest rates of infection were found in southern highbush `Georgiagem' (2.5%), `Misty' (2.5%), rabbiteye `Brightwell' (0.0%), and northern highbush `Bluecrop' (2.5%). Since many southern cultivars were infected by the disease, resistance likely has not excluded BBSSV from the southern United States.

Stability of PRV Resistance in Transgenic Papaya

Transgenic papaya lines carrying the coat protein gene (CP) of papaya ringspot virus (PRV) strain HA 5-1 display PRV reactions ranging from complete susceptibility (39-3 & 39-4), to slight delay in onset of symptoms (39-1) and attenuation of symptoms (60-3), to high-level resistance (55-1, 63-1). Normal Mendelian segregation of transgene expression was lost in R1 of 39-3 and 39-4, and inbred R1 60-3 gave an aberrant 1:1 ratio. R0 55-1 plants were resistant in the field (Hawaii) for 2 years following manual and/or aphid inoculation, and the high-level resistance remained stable in the R1 after repeated manual inoculations in the greenhouse and graft inoculation for up to 1 year (Cornell). However, inoculation with PRV HA-Oahu strain produced symptoms in some plants at Cornell (9% after 6 weeks) and in Hawaii (50% after 1 year). Two 55-1 and one 60-3 plant subsequently underwent remission of symptoms and became ELISA-negative (Hawaii). Transmission of PRV isolates from symptomatic 55-1 plants to other CP+ 55-1 bioassay plants was unsuccessful.

Resistance to Peanut Mottle Virus in Arachis spp.1

Seven wild rhizomatous peanut introductions [PI 262794, PI 262818, AM 3867, ‘Florigraze’ (PI 421707), PI 172223, ‘Arbrook’ (PI 262817), and ‘Arblick’ (PI 262839)] from the Plant Materials Center, Americus, Ga. were not infected with peanut mottle virus (PMV) by mechanical inoculation, aphid inoculation, or by natural infection when field planted near infected Arachis hypogaea L. These accessions are the only known sources of resistance to PMV in Arachis.