Oligochitosan induces cell death and hydrogen peroxide accumulation in tobacco suspension cells

Oligochitosan has been shown to induce several plant defense responses. In the present work, the effect of oligochitosan on tobacco cell survival was investigated. The results showed that oligochitosan caused tobacco cell death in a dose-dependent manner. About 40.6 % tobacco cells died when cultured for 72 h after 500 μg ml⁻¹ oligochitosan treatment. Certain aspects of this cell death process appeared to be similar to apoptosis in animal cells. These included shrinkage of cytoplasm and condensation of chromatin. Oligochitosan also induced H₂O₂ accumulation in tobacco cell suspension culture. The role of H₂O₂ in the signal transduction that leads to cell death was investigated. Co-treatment of tobacco cells with oligochitosan and catalase inhibited H₂O₂ accumulation but did not inhibit the induction of cell death. The results suggested that apoptosis-like cell death of tobacco cells induced by oligochitosan is independent of H₂O₂ signal pathway.     

Involvement of nitric oxide generation in hypersensitive cell death induced by elicitin in tobacco cell suspension culture

Recent studies suggest that nitric oxide (NO), an important signaling and defense molecule in mammals, plays a key role in activating disease resistance in plants. We characterized NO production by tobacco Bright Yellow-2 cells pharmacologically after treatment with INF1, the major elicitin secreted by the late blight pathogen Phytophthora infestans, prepared from Escherichia coli. NO production rapidly occurred within 1h and reached a maximum level 3–6h after the addition of INF1. Carboxy-PTIO, a NO-specific scavenger, abolished INF1-induced NO production in a dose-dependent manner. Pretreatment of protein synthesis inhibitor cycloheximide and protein kinase inhibitor K252a blocked NO production 3–12h after INF1 treatment, indicating that NO production requires de novo protein synthesis and protein phosphorylation. In an investigation of the relations between NO generation and several defense responses induced by INF1, carboxy-PTIO completely suppressed activation of a 41-kDa protein kinase and cell death by INF1. Carboxy-PTIO also suppressed the induction of hypersensitive-related (hsr) genes HSR515 and HSR203J, the expression of which is strongly correlated with the hypersensitive response in plants. The results suggest that NO plays a crucial role in the induction of hypersensitive cell death.     

电化学酶联免疫分析检测烟草花叶病毒和烟草环斑病毒

以邻苯二胺 (OPD)底物为例 ,用抗原直接包被法 (DAC -ELISA)同线性扫描极谱法 (LSV)相结合 ,检测烟草花叶病毒 (TMV)的提纯液 ,检出限可达 0 2 5ng/ml,TMV粗提液的最高稀释比为 1 :1 0 2 4 0 0 ;检测烟草环斑病毒 (TRSV)提纯液 ,检出限为 1 0ng/ml,粗提液的最高稀释比为 1 :1 0 0 0 0 ,该方法的灵敏度比DAC -ELISA光度法提高了 5倍以上     

Tobamoviruses of pepper,eggplant and tobacco: Comparative host reactions and serological relationships

Using test plants and serology six tobamoviruses of pepper (FO, Ob, P8, P11, P14 and SL) and one of eggplant (A1) were compared with common tobacco mosaic virus (TMV-WU1). WU1, A1 and FO were closely similar in their reactions inCapsicum spp. as were P14 and SL. Ob, P11 and P8 were also similar in this respect except inC. frutescens Tabasco in which P8 differed from Ob and P11.Using micro-precipitation tests the virus strains could be roughly divided into three serological groups: Group I consisted of WU1, group II of A1, FO, P8, P14 and SL, and group III of P11 and Ob. With ELISA group II was further divisible into two subgroups, including A1 and FO, and P8, P14 and SL.It was concluded that similarities of strains in their reactions inCapsicum spp., were not necessarily confirmed by their serological relationships.Samenvatting Zes tobamovirussen uit peper (FO, Ob, P8, P11, P14 en SL) en één uit aubergine (A1) konden met behulp van toetsplanten alle van elkaar worden onderscheiden. In hun reacties inCapsicum-soorten, kwamen A1 en FO sterk overeen met elkaar en met het gewone tabaksmozaïekvirus (WU1). Ob, P11 en P8, die in dit opzicht onderling veel overeenkomst vertoonden, verschilden duidelijk van alle andere. Hetzelfde gold voor P14 en SL.Ook met behulp van de micro-precipitatietoets konden de virusstammen in groepen worden ingedeeld. Groep I werd gevormd door WU1, groep II door A1, FO, P8, P14 en SL en groep III door P11 en Ob. Met behulp van ELISA kon groep II worden onderverdeeld in twee ondergroepen, namelijk A1 en FO enerzijds en P8, P14 en SL anderzijds.De nauwe serologische verwantschap van A1 met FO is conform de grote overeenkomst in waardplantreacties. Hetzelfde geldt voor P11 en Ob, wanneer we alleen hun reacties inCapsicum-soorten beschouwen. P8 echter, die wat het laatste betreft meer op Ob en P11 lijkt, vertoonde serologisch meer overeenkomst met P14 en SL. WU1 verschilde serologisch zeer sterk van alle andere onderzochte virusstammen.Geoconcludeerd kan worden dat de waargenomen overeenkomst tussen de onderzochte virusstammen in hun reacties inCapsicum-soorten niet altijd gesteund wordt door hun serologische verwantschappen.Guestworker from September 1981 till January 1982 as a fellow of the International Agricultural Centre, Wageningen, the Netherlands, from the Vegetable Crops Research Institute, Budapest, Hungary.Seconded to the Glasshouse Crops Research and Experiment Station, Naaldwijk, the Netherlands.     

Comparison of the effects of salicylic acid and ethephon with virus-induced hypersensitivity and acquired resistance in tobacco

The induction of a hypersensitive reaction in Samsun NN tobacco by tobacco mosaic virus (TMV) at 20°C leads to the development of both localized and systemic acquired resistance, and is associated with the appearance of pathogenesis-related proteins (PR's) and large increases in peroxidase activity and ethylene production. Salicylic acid (SA) induced a similar resistance in treated plant parts and occasionally also in untreated upper leaves of plants of which three lower leaves had been injected. SA also induced the same four PR's, but these were confined to the treated leaves. Thus, the connection between the presence of PR's and the reduction of TMV multiplication and spread may not be direct.In contrast to TMV, SA did not stimulate ethylene production and hardly increased peroxidase activity. Induction of acquired resistance and PR's by SA developed equally well at 20°C and at 32°C. However, pricking leaves with needles moistened with the ethylene-releasing compound ethephon mimicked TMV infection in inducing acquired resistance and PR's in both treated and untreated leaves at 20°C, but not at 32°C. Ethephon increased peroxidase activity at both temperatures, but only at 20°C dit it induce changes in both the anodic and the cathodic isoenzymes that were similar to those induced as a result of TMV infection. SA induced PR's and reduced TMV multiplication in Samsun tobacco, and inhibited virus spread in Samsun NN at 32°C.These observation indicate that neither the induction of PR's, nor the development of acquired resistance is temperature-sensitive. On the other hand, the effects of ethephon are temperature-sensitive in the same way as the hypersensitive response to TMV. It can thus be hypothesized that ethylene, produced naturally during the hypersensitive reaction of tobacco to TMV, leads to the temperature-sensitive synthesis or release of a presumably benzoic acid-type compound that functions as the natural inducer of PR's and acquired resistance. Although vanillic acid has been shown to accumulate in hypersensitively reacting tobacco leaves, it produced none of the effects of SA, and thus cannot be the natural inducer.Samenvatting Inductie van een overgevoeligheidsreactie in Samsun NN-tabak door tabaksmozaïekvirus (TMV) bij 20°C leidt tot de ontwikkeling van een verworven resistentie die zowel lokaal als systemisch werkzaam is, en gaat samen met het verschijnen van pathogenesis-related proteins (PR's) en sterke toename in de activitieit van peroxidase en de produktie van ethyleen. Salicylzuur (SA) induceerde een vergelijkbare resistentie in behandelde plantedelen en af en toe ook in niet behandelde bovenbladeren van planten waarvan drie onderbladeren waren ingespoten. SA induceerde ook dezelfde vier PR's, maar deze waren beperkt tot de behandelde bladeren. Er bestaat dus geen directe samenhang tussen de aanwezigheid van PR's en de remming van de vermeerdering en uitbreiding van TMV in de plant.In tegenstelling tot TMV stimuleerde SA de ethyleenproduktie niet en verhoogde het de peroxidaseactiviteit nauwelijks. Inductie van verworven resistentie en PR's door SA trad even goed op bij 32°C als bij 20°C. Net als infectie met TMV leidde aanprikken van bladeren met naalden die gedoopt waren in een oplossing van ethefon — waaruit in het blad ethyleen vrijkomt — echter tot inductie van verworven resistentie en PR's in zowel behandelde als onbehandelde bladeren bij 20°C, maar niet bij 32°C. Ethefon verhoogde de peroxidaseactiviteit bij beide temperaturen, maar alleen bij 20°C induceerde het veranderingen in zowel de anodische als de kathodische isoënzymen die vergelijkbaar waren met die welke geïnduceerd werden als gevolg van infectie met TMV. SA induceerde PR's en verminderde de vermenigvuldiging van TMV in Samsun tabak, en remde de uitbreiding van het virus in Samsun NN bij 32°C.Deze waarnemingen tonen dat noch de inductie van PR's, noch de ontwikkeling van verworven resistentie een temperatuurgevoelig proces is. Daarentegen zijn de effecten van ethefon op dezelfde wijze temperatuurgevoelig als de overgevoeligheidsreactie op TMV. Men kan daarom veronderstellen dat ethyleen, dat op natuurlijke wijze geproduceerd wordt tijdens de overgevoeligheidsreactie van tabak op TMV, aanleiding geeft tot een temperatuurgevoelig proces, namelijk de synthese of het vrijkomen van een verbinding, vermoedelijk een benzoëzuurderivaat, dat fungeert als de natuurlijke inductor van PR's en verworven resistentie. Hoewel is aangetoond dat vanillinezuur zich ophoopt in overgevoelig reagerende tabaksbladeren, veroorzaakte deze verbinding geen enkel van de effecten van SA. Vanillinezuur kan dus niet de natuurlijke inductor zijn.     

检测兰花病毒的斑点酶联法的建立

本文建立了齿兰环斑病毒（Odntoglossum ringspot virus,ORSV）和建兰花叶病毒（Cymbidium mosaic virus,CyMV）两种兰花主要病毒的斑点酶联法检测技术（Dot-ELISA）。用Dot-ELISA对提纯病毒和田间病叶进行检测,结果表明;检测ORSV和CyMV提纯病毒的灵敏度分别为0.82μg/mL和0.244μg/mL;病叶汁液的稀释倍数分别为1280倍和2650倍。该方法具有操作简便、成本低等优点,适合于兰花生产中的病毒检测。     

利用黄瓜花叶病毒M和Fny株系 的假重组体分析致病关键因子

 用RT-PCR扩增黄瓜花叶病毒M株系（CMV-M）全长基因组cDNA,成功构建CMV-M RNA2和RNA3侵染性克隆后,与CMV-Fny基因组RNA交换得到3个假重组型病毒 （F1M2F3、F1F2M3、F1M2M3）。用F1M2F3、F1F2M3、F1M2M3分别侵染白肋烟,产生坏死环斑、轻微绿斑驳、明脉、黄白化和叶尖线性化等症状。根据假重组型病毒和野生型病毒的表观症状,分析引起各种症状的关键因子,初步判定：CP基因是诱导花叶症状的关键因子,CMV-Fny RNA2是诱导叶尖线性化的关键因子,CMV-M RNA2是诱导叶尖坏死斑关键因子。实时荧光定量PCR结果显示：野生株CMV-M、CMV-Fny和假重组体F1M2F3、F1F2M3、F1M2M3侵染烟草后引起的症状差异与病毒基因组RNA累积没有直接关系。     

Interaction between the helicase domain of the tobacco mosaic virus replicase and a tobacco arginine decarboxylase

In a yeast two-hybrid screening test for tobacco proteins that interact with TMV replicase using the helicase (H) domain as bait, a cDNA clone was selected that encodes a polyamine biosynthetic enzyme, arginine decarboxylase (ADC). In yeast cells, the C-terminal internal region of ADC interacted with the H domain. This observation was confirmed in vitro by far-Western blotting. Inhibition of the binding between the H domain and the IRnHEL (I region and N-terminus of helicase domain) region by ADC using a yeast three-hybrid assay suggested possible interference of the heterodimerization of 126K and 183K by ADC.The nucleotide sequence data of pADCF reported in this study is available in the DDBJ/EMBL/GenBank databases under accession number AB110952     

Pelargonium flower-break and pelargonium line pattern viruses in the Netherlands; purification,antiserum preparation,serological identification,and detection in pelargonium by ELISA

Two viruses, detected frequently in the Netherlands in pelargonium, were identified by serology and test plant reactions. Antisera were prepared and an ELISA procedure was developed to detect the viruses in pelargonium.One of the viruses, PFBV-N, proved to be pelargonium flower-break virus. With the antiserum to PFBV-N, it could be detected reliably throughout the year inPelargonium zonale Springtime Irene.The other virus, PLPV-N, was serologically closely related to pelargonium line pattern virus (PLPV) and to pelargonium ring pattern virus (PRPV), as were an old virus isolate from Saturnus, collected in the Netherlands in 1971 (L128), and PLPV isolates from Yugoslavia (PLPV-Y) and Denmark (PLPV-D). There were only minor differences in host-plant reactions between the virus isolates. Based on these tests, PLPV and PRPV are considered as isolates of the same virus, for which, for practical reasons, the name pelargonium line pattern virus is proposed.PLPV could be reliably detected by ELISA inP. zonale Springtime Irene and Amanda throughout the year with only a few exceptions. InPelargonium peltatum Tavira, however, reslts were erratic due to uneven distribution of virus in the plant. Best results were obtained when petioles of fully expanded leaves were tested.     

Characterisation of BaYMV and BaMMV Pathotypes in France

The reaction of thirty-four barley cultivars from European and Asiatic origin was analysed in six soils infected with barley yellow mosaic virus complex (BaYMV, BaMMV). These soils were selected from 16 sites for their differences in cultivar response. Amongst the six cultivars carrying the ym4 gene (Esterel, Express, Labéa, Majestic, Réjane, Vanoise), only cv Majestic was infected at one site with BaYMV and BaMMV. Concerning BaYMV, three cultivars were infected on all the soils and 19 on none of them. Twelve cultivars were differentially infected depending on the soil. In the case of BaMMV, four cultivars were infected on all the soils and 19 on none of them. Eleven cultivars were differentially infected depending on the soil. ELISA tests revealed the presence, in these soils, of variants of BaYMV and BaMMV that were able to overcome at least seven of the 12 known resistance genes (ym3, ym4, ym6, ym8, ym9, ym10, ym11) and the resistance of three varieties (Tosan Kawa 73, OU1 and Taihoku A) in which the genetic basis is unknown. Amplification by RT-PCR of the N-terminal region for three of BaYMV variants was performed. Nucleotide and amino acid sequences were determined and compared with the corresponding sequence of a common strain of BaYMV-G. A few nucleotide differences were detected between all the French isolates, but there were no strain specific amino acid differences.     

Shoot meristem tissue of tobacco inoculated with Cucumber mosaic virus is infected with the virus and subsequently recovers from infection by RNA silencing

Distribution of Cucumber mosaic virus (CMV) in shoot meristem tissue of CMV-inoculated tobacco was successively analyzed with immunohistochemical microscopy and in situ hybridization. CMV signals were detected in the tissue at 7 days postinoculation (dpi), but then they decreased and disappeared after 14dpi. Detailed observation confirmed CMV invasion of shoot apical meristem at 6–8dpi. Short interfering RNA corresponding to CMV RNAs was first detected at 7dpi and was detected up to 24dpi. These results suggest that the shoot meristem tissue is infected with CMV but subsequently recovers from the infection by RNA silencing.     

The long-term preservation of potato virus Y and watermelon mosaic virus in liquid nitrogen in comparison to other preservation methods

Samples of watermelon mosaic virus (WMMV), consisting of clarified sap of diseased squash plants, were found to be very infective after more than 4 years of storage if the samples had been deep frozen at –18°C, and to have lost most of their infectivity within 4 years of preservation by freeze drying and subsequent storage of the samples at 4°C.Potato virus Y (PVY) inocula, consisting of samples of clarified sap of diseased red peppers, lost their infectivity in less than 4 years if the samples had been deep frozen at –18°C but retained their infectivity for more than 4 years if the samples had been freeze dried and stored at 4°C.A decrease in infectivity of both viruses in the deep frozen at –18°C and freeze dried inocula could be observed even during the first months of storage. However, samples of clarified sap stored in or over liquid nitrogen maintained their activity for at least 22 months for WMMV and 32 months for PVY with no indication of a decrease in infectivity of the viruses. Storage in or over liquid nitrogen seems therefore a very promising long-term preservation method for plant viruses.Samenvatting Watermeloenmozaïekvirus inocula, bestaande uit helder gecentrifugeerd sap van zieke squashplanten, bleken hun infectievermogen zeer goed te hebben behouden na meer dan vier jaar bewaring bij –18°C, maar binnen vier jaar grotendeels te hebben verloren na droogvriezen en bewaren bij 4°C.Overeenkomstig bereide aardappelvirus Y inocula uit zieke paprikaplanten hadden hun infectievermogen binnen vier jaar verloren na bewaring bij –18°C, maar konden hun infectievermogen grotendeels behouden gedurende meer dan vier jaar na droogvriezen en bewaring bij 4°C.Binnen enkele maanden na het begin der bewaringsperiode kon al een afname van het infectievermogen van beide virussen in de diepgevroren (–18°C) en drooggevroren inocula worden gevonden. Helder gecentrifugeerde sapmonsters die in of boven vloeibare stikstof bewaard waren, hadden gedurende tenminste 22 maanden voor het watermeloenmozaïekvirus en 32 maanden voor het aardappelvirus Y hun volle infectievermogen behouden. Bewaring in of boven vloeibare stikstof is daarom de beste van de drie getoetste bewaarmethodes voor opslag op lange termijn van beide virussen. Deze methode lijkt ook veelbelovend voor een groot aantal andere virussen.     

Occurrence of barley yellow mosaic and barley mild mosaic bymoviruses in Greece

In March 1991, large chlorotic patches appeared in an autumn-sown barley crop growing near Thessaloniki, Greece. Leaves had characteristic mosaic symptoms and immunosorbent electron microscopy and enzyme-linked immunosorbent assay confirmed the presence of both soil-borne mosaic viruses of barley, barley mild mosaic and barley yellow mosaic bymoviruses. In the following year, similar symptoms appeared in a crop at Souroti, 30 km east of Thessaloniki but the disease has not been found in other areas of Macedonia. This report is the first record of these viruses from Greece and is the most southerly European record.     

Induction of a viable but not culturable (VBNC) state in some Pseudomonas syringae pathovars upon exposure to oxidation of an apoplastic phenolic,acetosyringone

Acetosyringone is a phenolic metabolite often found in plant apoplasts. Its oxidation by hydrogen peroxide and peroxidase results in a prolonged increase in the redox potential of the reaction mixture, similar to redox increases observed in tobacco suspension cells upon treatment with incompatible bacteria. Since high redox potentials, being oxidative, are generally detrimental to bacteria, the effect of acetosyringone oxidation on bacterial viability was examined. Pseudomonas syringae pv. syringae was added to reaction mixtures containing acetosyringone, hydrogen peroxide and peroxidase and samples were removed to determine viability by dilution plating. Initial studies were done with low bacterial concentrations, 10⁵ CFU ml⁻¹, to ensure that scavenging of H₂O₂ was negligible and did not interfere with the reaction mixture. No colonies were formed by bacteria that had been added to reaction mixtures with acetosyringone ranging from 25 to 100 μΜ. Examination of the bacteria by microscopy and flow cytometry, using fluorescent stains that indicate bacterial membrane integrity, suggested that these bacteria had maintained their membrane integrity. In addition they were able to respire based on oxygen uptake. When bacteria were added to on-going reaction mixtures at a time point after the prolonged redox response, the CFU ml⁻¹ increased indicating that a stable reaction product was not responsible for the non-culturability bioactive effect. Other bacterial isolates, P. s. pv. tabaci and Pseudomonas fluorescens, were less susceptible to the bioactive effect of the acetosyringone oxidation. Other phenolics were tested and had lesser degrees of bioactivity and in some cases reduced the bioactivity of acetosyringone oxidation. The ‘viable but non-culturable’ (VBNC) state of the bacteria in this study is compared to that described for other medical and plant pathogens.     

Amino acid 129 in the coat protein of Cucumber mosaic virus primarily determines invasion of the shoot apical meristem of tobacco plants

We previously reported that a strain of Cucumber mosaic virus (Pepo CMV) invaded the shoot apical meristem (SAM, tunica corpus) of tobacco plants at 6–8 days postinoculation (dpi), contrary to earlier observations. To identify a viral factor determining the ability to invade the SAM, we inoculated plants with two other CMV strains, MY17 and Y, and tested the three strains in this study. Immunohistochemical microscopy revealed that MY17 CMV invaded the SAM at 7 dpi, the same as Pepo CMV, but Y CMV did not, even at 21 dpi. Using RNA pseudorecombinants between Pepo and Y CMV, we found that Pepo RNA 2 affected the rate of SAM invasion, and Pepo RNA 3 was required for successful SAM invasion. Inoculation with RNA 1 and RNA 2 from Y CMV and RNA 3 containing the chimeric coat protein (CP) gene between Pepo and Y CMV or a Y RNA 3 point mutant containing a Ser-to-Pro substitution at position 129 in CP (Y129P) revealed that amino acid 129 of CP is the determinant for successful SAM invasion. The rate of SAM invasion of the pseudorecombinants and Y129P was consistent with the efficiency of cell-to-cell movement in the inoculated leaves, implying that SAM invasion by CMV strains may be due to efficient cell-to-cell movement.     

Acquired resistance triggered by elicitins in tobacco and other plants

Elicitins are a family of proteins excreted byPhytophthora spp. They exhibit high sequence homology but large net charge differences. They induce necrosis in tobacco plants which then become resistant to the tobacco pathogenPhytophthora parasitica var.nicotianae. In stem-treated plants, resistance was not restricted to the site of elicitin application, but could be demonstrated by petiole inoculation at all levels on the stem. Resistance was already maximum after two days and lasted for at least two weeks. It was effective not only towardsP. p. var.nicotianae infection, but also against the unrelated pathogenSclerotinia sclerotiorum. In contrast to dichloroisonicotinic acid, an artificial inducer of systemic acquired resistance, which was increasingly effective with doses ranging from 0.25 to 5mole per plant, the basic elicitin cryptogein exhibited a threshold effect, inducing near total resistance and extensive leaf necrosis above 0.1 nmole per plant. Between 1 and 5 nmole, acidic elicitins (capsicein and parasiticein) protected tobacco plants with hardly any necrotic symptom. Elicitins exhibited similar effects in various tobacco cultivars andNicotiana species, although with quantitative differences, but induced neither necrosis nor protection in other SolanaceÆ (tomato, petunia and pepper). Among 24 additional species tested belonging to 18 botanical families, only some BrassicaceÆ, noticeably rape, exhibited symptoms in response to elicitins, in a cultivar-specific manner. Elicitins appear to be natural specific triggers for systemic acquired resistance and provide a tool for unraveling the mechanisms leading to its establishment.Abbreviations AR acquired resistance - HR hypersensitive response - INA 2,6-dichloroisonicotinic acid - Ppn Phytophthora parasitica var.nicotianae - SAR systemic acquired resistance     

Mechanism of resistance to spinosyn in the tobacco budworm, Heliothis virescens

Topical laboratory selection of tobacco budworm larvae, Heliothis virescens, with technical spinosad for multiple generations resulted in larvae 1068-fold resistant to topical applications of the insecticide and 316.6-fold resistant to insecticide treated diet as compared to the parental strain. The penetration of 2′-O-methyl[¹⁴C]spinosyn A across the cuticle of the susceptible (parental) and selected (resistant) tobacco budworms increased with time 3-12 h after application. A trend of reduced penetration in the resistant strain was found but the differences were not statistically significant. 2′-O-methyl[¹⁴C]spinosyn A when injected into the hemocoel was not metabolized 96 h after treatment in both the susceptible and resistant strain, suggesting that a change in metabolism was not the mechanism of resistance. Electrophysiological studies indicated that dose-dependent spinosyn A-induced currents occurred in neurons from spinosyn resistant and susceptible (adult) tobacco budworms. At both 10 and 100 nM spinosyn A, however, the amplitude of these currents in the resistant insects was significantly smaller than the amplitude of currents observed from neurons from susceptible tobacco budworm adults. This suggests that neurons from resistant insects have decreased sensitivity to spinosyn A. However, the reduced inward currents in the resistant strain may or may not be related to the mode of action of the spinosyns. No statistically significant cross-resistance was noted for the spinosad resistant tobacco budworms for topical applications of permethrin (Pounce^®), profenofos (Curacron^®), emamectin benzoate (Denim^®), or indoxacarb (Steward^®). A statistically significant reduction in susceptibility to acetamiprid (Mospilan^®) in artificial diet as determined from a resistance ratio of 0.482 was found.     

Involvement of NbNOA1 in NO production and defense responses in INF1-treated Nicotiana benthamiana

During defense responses, plant cells produce nitric oxide (NO), which may control many physiological processes. In a previous study, we reported that nitrate reductase (NR) is responsible in part for INF1 elicitor-induced NO production in Nicotiana benthamiana, but the possibility remains that other NO-generating system(s) contribute to NO production. In mammalian cells, NO production is catalyzed by NO synthase (NOS). However, NOS-like enzyme(s) have never been identified in plants, and only the gene for Arabidopsis thaliana nitric oxide-associated 1 (AtNOA1) has been identified as a putative regulator of NOS activity in plants. In this study, we cloned NbNOA1, a homolog of AtNOA1, from N. benthamiana and investigated its involvement in NO production induced by INF1. The NbNOA1 gene was silenced by a virus-induced gene-silencing (VIGS) technique. NbNOA1-silenced plants had yellowish leaves. Silencing NbNOA1 partially decreased INF1-induced NO production, while overexpressing NbNOA1 did not affect NO production. Silencing NbNOA1 suppressed INF1-induced PR1a gene expression and increased susceptibility to Colletotrichum lagenarium. These results suggest that NbNOA1 is involved in INF1-mediated NO production and is required for defense responses. The nucleotide sequence data reported are available in the DDBJ/EMBL/GenBank databases under accession number AB303300.     

Species composition and abundance of aphids in Australian hop gardens and their impact on spatiotemporal patterns of Carlavirus epidemics

The species composition and abundance of aphids in commercial cv. Agate and cv. Super Pride hop gardens in Tasmania, Australia, were characterized over three seasons (1999–2001). Gunns Plains recorded 14 aphid species and Bushy Park 11 species, with nine of these common to both sites. The majority of aphids were trapped in the first 2 months (October and November) of active hop growth in all three seasons. Cultivar and geographical location had significant effects on the abundance of total aphids (species pooled) trapped and several individual aphid species in the three seasons. In general, significantly more aphids (total and individual species) were trapped in cv. Agate than cv. Super Pride gardens, and higher numbers were trapped at Bushy Park than at Gunns Plains. This coincided with a higher incidence of plants infected by carlaviruses in cv. Agate gardens at both locations. Differences in the spatiotemporal dynamics of Carlavirus epidemics were described by fitting a stochastic model to the data, with parameters for local spread within the garden (contagion) and background infection (disease increase unrelated to infected plants within the gardens). Local spread of Hop latent virus (HpLV) and Hop mosaic virus (HpMV) was indicated within all gardens. For HpMV in cv. Agate at Gunns Plains, however, infections caused by immigrant viruliferous aphids were also apparent. Using join-count statistics, spatial aggregation of both virus diseases was found for all years, except for the initial year (1999) when incidence was low. Clusters of diseased plants extended to greater distances for HpLV than for HpMV. Based on spatial and spatiotemporal analyses, local spread (mechanical transmission and/or aphid movement within the garden) appears to be the dominant factor in the epidemics of HpLV. Aphid immigration from outside the crop over time may play a more significant role for HpMV epidemics, at least for one location.