Effects of light quality on conidiophore formation of the melon powdery mildew pathogen <Emphasis Type="Italic">Podosphaera xanthii</Emphasis>

The lengths of conidiophores in fungal colonies of the melon powdery mildew pathogen Podosphaera xanthii Pollacci KMP-6 N cultured under greenhouse (natural) conditions differed markedly from those cultured in a growth chamber. We hypothesized that light wavelength was responsible for the differences in conidiophore length. In this study, we examined the effects of light-emitting diode (LED) irradiation (purple, blue, green, orange, and red light) and white light on colony development and conidiophore formation in KMP-6 N using a stereomicroscope and a high-fidelity digital microscope. Colonies on leaves were flat under greenhouse conditions and under red LED light irradiation but were stacked under growth chamber conditions and under purple, blue, green, and orange LED light irradiation. In addition, KMP-6 N formed catenated conidia comprising six conidia per conidiophore under greenhouse conditions and red light but more than seven conidia per conidiophore under growth chamber conditions and purple, blue, green, and orange light. Furthermore, almost none of the conidia on top of the conidiophores grown under blue light were fully constricted. Therefore, these fungi could not scatter their conidia and spread infection. This is the first report of the effects of LED lights on conidiophore formation in the melon powdery mildew fungus P. xanthii. The results provide insight into the mechanisms underlying the responses of conidiophores to light of specific wavelengths and conidial scatter from conidiophores of melon powdery mildew fungi.     

Infection process of <Emphasis Type="Italic">Pseudocercospora musae</Emphasis> on banana leaf

Yellow Sigatoka that is caused by Pseudocercospora musae is an important banana disease. The aim of this study was to elucidate the infection process of P. musae in banana leaves by scanning electron microscopy. Leaf samples were inoculated on the abaxial surface with P. musae and then analysed at 12, 24, 36, 48, 72, 96, 120, 144, and 168 h post inoculation (hpi) and at 36 and 50 days post inoculation (dpi). The conidia were found to be germinated between 24 and 36 hpi and penetrated through the stomata between 96 and 120 hpi, or more generally from 144 hpi. P. musae colonized the spongy parenchyma at 36 dpi and the palisade parenchyma at 50 dpi. Sporulation occurred at 50 dpi on the adaxial surface of leaves through the emergence of conidia on conidiophores through the stomata. Considering the importance of yellow Sigatoka in banana production, our results provide a better understanding of the life cycle of the fungus for treatment processes.     

Resistance levels of cucumber to <Emphasis Type="Italic">Podosphaera xanthii</Emphasis> in a growth chamber are related to haustorial formation and hyphal branching frequency

To clarify relationships between powdery mildew resistance of cucumber and hyphal developmental state of the pathogen, haustorial formation and the hyphal branching frequency were compared among cucumber varieties that differ in resistance levels to powdery mildew pathogen Podosphaera xanthii. Cotyledons of four cultivars were inoculated with P. xanthii. By 2 days after inoculation, secondary haustoria had developed from the first hyphal cell that formed beside the conidium in susceptible cultivars. The fungus on susceptible cultivars also tended to have hyphal branches just after the hyphal cell producing haustoria.     

Biological control mechanisms of D-pinitol against powdery mildew in cucumber

D-pinitol is an effective agent for controlling powdery mildew (Podosphaera xanthii) in cucumber. In this study, we determined the mechanisms of D-pinitol in controlling powdery mildew in cucumber plants. We compared P. xanthii development on cucumber leaf surface treated with D-pinitol or water (2 mg ml⁻¹) at different time points after inoculation. The germinating conidia, hyphae, and conidiophores of the pathogen were severely damaged by D-pinitol at any time of application tested. The highest level of suppression of fungal development was obtained at 3 days after inoculation. The contents of chlorophyll, total phenolics, flavonoid, and gallic acid and its derivatives (GAD); the activities of phenylalanine ammonialyase (PAL), polyphenoloxidase (PPO), peroxidase (POX), and superoxide dismutase (SOD); and the expression of the genes encoding for PR-1, peroxidase (POX), lipoxygenase (LOX1), chitinase (Chit1) were higher in the cucumber leaves treated with D-pinitol and inoculated than in the leaves either treated with D-pinitol or inoculated with the pathogen. These results suggest that D-pinitol triggers several plant defense responses in cucumber leading to pathogen suppression and resistance to powdery mildew.     

Erarbeitung eines Testverfahrens zur Bestimmung der Protektiv- und Kurativleistung von Fungiziden bei der Bek?mpfung des Erregers der Stemphylium-Laubkrankheit Stemphylium botryosum in der Spargelkultur (Asparagus officinalis L.)

The foilage of Asparagus officinalis L. has a large contact surface for different leaf diseases, which can lead to profits cuts, especially by the causal pathogen of Stemphylium leaf spot, Stemphylium botryosum. The maintaining healthy of the foliage is an important precondition to achieve an optimal harvest in the following year. Therefore, it is necessary to describe the efficacy of currently approved plant protection products against S. botryosum in order to optimize the fungicide applications. In the context of greenhouse trials with asparagus seedlings a method should be developed to evaluate the preventive and curative efficacy of fungicides. At the beginning, stable, repeatable and homogeneous infections of S. botryosum needed to be achieved. Ad hoc the factors plant age (weeks after sowing date), inoculum (density of conidia per ml and concentration of malt), infection conditions (period of high relative humidity) and day of rating, as well as their influence on the occurrence of the symptoms, were investigated. With the exception of the factor plant age, the other factors had a significant effect on the infection. For the trials to evaluate the efficacy of fungicides, the analysed factors were defined. The age of the seedlings was six weeks after sowing, the inoculum was composed of 1,00?×?10⁵ conidia per ml combined with an 0,5% malt concentration, as infection condition the ??tunnel system?? with periodically high relative humidity was chosen. 13 days past infection was the optimal day of rating. To apply the seedlings with the fungicides, the registered application rate was reduced to 1/5. The trials to evaluate fungicides were conducted with the fungicides Signum^®, Rovral^® WG and two different testing agencies. At four consecutive application days, the plants were covered with fungicides. In the following, the infestation degree of stems in percent was evaluated. Additionally, the used S. botryosum-Isolates were biomolecularly investigated and the QoI- and SDHI resistance state determined.     

Production of <Emphasis Type="Italic">Guignardia bidwellii</Emphasis> conidia on grape leaf lesions is influenced by repeated washing events and by alternation of dry and wet periods

The ascomycete Guignardia bidwellii is an economically important pathogen in many grapevine-growing areas. Primary infections are caused by ascospores and conidia produced in mummified berries and in cane lesions. Secondary infections are caused by the conidia produced by pycnidia formed in leaf lesions and, in later season, in rotted berries. Environment-controlled experiments were conducted to study the production dynamics of G. bidwellii conidia on grape leaf lesions as influenced by: i) repeated washing events, and ii) alternate dry and wet periods. Under optimal environmental conditions (25 °C, 100 % relative humidity), production of conidia declined over washings and was almost completely depleted after four washings. When pycnidia were kept in a low humidity environment (average of 54 % relative humidity) between two successive washings, the production of conidia progressively diminished as the time between washings increased, with few conidia being still produced after 87 days. This decline in conidial production was faster at 29 °C than at 20 °C. This information is relevant in that it determines the potential of black-rot lesions to produce conidia along the grape-growing season and, therefore, their contribution to epidemic development.     

Development of tolerance to the diphenyl ether herbicide acifluorfen-methyl in excised cucumber (Cucumis sativus L.) cotyledons

Etiolated cucumber (Cucumis sativus L.) cotyledons and cotyledons greened for 24 hr in low light (75 μE m⁻² sec⁻¹; measured as PAR, i.e., photosynthetically active radiation between 400 and 700 nm) were susceptible to 1 μM acifluorfen-methyl (AFM), methyl-5-[2-chloro-4-(trifluoromethyl)phenoxy]-2-nitrobenzoate, following an 8-hr exposure to high light (600 μE m⁻² sec⁻¹; PAR). Cotyledons greened in low light for 48 hr prior to treatment with 1 μM AFM in high light were tolerant. Injury was detected by monitoring efflux of 3-O-methyl-[¹⁴C]glucose from treated cotyledons excised from 6- or 7-day-old dark-grown seedlings. Although development of tolerance was light dependent, tolerance was not related specifically to seedling age. Tolerance to AFM injury was overcome partially by rubbing the adaxial surface of the leaf (i.e., the side of the cotyledon exposed to herbicide) with a wetted finger and was eliminated completely following abrasion with carborundum. Cuticle abrasion resulted in an increase in [¹⁴C]AFM uptake. Scanning electron microscopy revealed that abrasion resulted in disruption of the leaf surface. These induced aberrations in surface structure facilitated enhanced absorption of AFM. Nonpolar hydrocarbon constituents of cuticular and epicuticular waxes of 24- and 48-hr-greened cotyledons were examined using gas chromatography. There were no differences in these cuticular components. Transmission electron micrographs indicated there were also no differences in cuticle thickness. The light-dependent development of tolerance to AFM activity was due in part to a decrease in herbicide absorption. The mechanism(s) responsible for inhibition of herbicide uptake and tolerance are unknown.     

Inoculum availability and seasonal survival of Potebniamyces pyri in pear orchards

Potebniamyces pyri (anamorph Phacidiopycnis piri) is the causal agent of Phacidiopycnis rot, a postharvest disease of pear fruit (Pyrus communis). Infections of pear fruit by P. pyri occur in the orchard, and symptoms develop after harvest during storage or in the market. P. pyri also is the cause of a canker and twig dieback disease of pear trees. To determine inoculum availability of P. pyri, dead bark and dead fruit spurs were periodically collected in two commercial ‘d’Anjou’ pear orchards and examined for the presence and viability of fruiting bodies of P. pyri. To determine seasonal survival of P. pyri, 2-year-old twigs of ‘d’Anjou’ pear in a research orchard were inoculated approximately monthly over 2 years with P. pyri and monitored for canker development. Inoculated twigs were removed from the trees 6 months post inoculation and examined for formation, viability of pycnidia of P. pyri, and reisolation of the pathogen. In both commercial orchards, all sampled trees were infected by P. pyri; viable pycnidia of P. pyri were observed on 42–78 % of the sampled bark and 5–32 % of the sampled fruit spurs; and viable conidia were observed at all sampling times during the fruit growing season. Apothecia of P. pyri also were observed on sampled dead bark and fruit spurs, but at a frequency ranging from 0 % to 19 %. P. pyri was recovered from inoculated twigs 6 months after inoculation at all sampling times during the 2-year study, but recovery frequency varied. P. pyri formed pycnidia on most cold-injured and inoculated twigs. The results suggest that: i) the conidial state of P. pyri is the main type of inoculum in pear orchards in the region; ii) viable inoculum of P. pyri for potential fruit infections is available during the pear fruit-growing season; iii) P. pyri can form pycnidia on cankers of twigs infected by the fungus at different seasons during the year; and iv) P. pyri can survive as mycelium in diseased pear twigs year round in the orchard.     

Biological characterization and genetic diversity analysis of two species of Pestalotiopsis causing twig dieback of Myrica rubra

Chinese bayberry (Myrica rubra Siebold & Zucc) is an evergreen fruit tree with high ecological and economic values in China. In recent years, a new twig dieback disease caused by Pestalotiopsis mangiferae and P. vismiae was observed in major M. rubra-producing areas of Zhejiang and Fujian Provinces, causing serious economic losses. In this study, 16 isolates of P. mangiferae and 27 isolates of P. vismiae were obtained from diseased leaves, roots and branches of M. rubra from different regions. The optimum growth temperature of the two species of Pestalotiopsis was determined to be 20–25 °C, while the optimum temperature for the germination of conidia was 25–35 °C. The two species of Pestalotiopsis showed rich genetic diversity. Inoculating the conidial suspension of one or both of the two species of the Pestalotiopsis on detached leaves or branches of M. rubra could cause lesions surrounding the inoculation sites with the frequency of 100 %. Moreover, necrotic lesions could be observed on inoculated potted plants with the frequencies of 33.3 % for P. mangiferae, 25 % for P. vismiae and 50 % for a mixed inoculum.     

Effect of temperature on infection and development of powdery mildew on cucumber

Podosphaera xanthii and Golovinomyces orontii are the causal agents of cucurbit powdery mildew. The effect of temperature on conidial germination, infection and sporulation was studied under controlled conditions. Conidia were inoculated on cucumber leaf discs, and incubated at six constant temperatures (from 10 to 35 °C in 5 °C steps) for 3 to 72 h to evaluate conidial germination and infection, and for 6–15 days to evaluate sporulation intensity. Germination took place at all tested temperatures, but was close to zero at 35 °C. The longest germ tubes measured in this experiment were 141.74 μm for the secondary germ tube of P. xanthii at 20 °C after 48 h of incubation, and 67.92 μm for G. orontii for the primary germ tube at 20 °C after 48 h of incubation. The optimal temperatures for conidial germination, infection and sporulation were 24.4, 25.7 and 22.3 °C, respectively, for P. xanthii, and 17.9, 17.3 and 14.9 °C, respectively, for G. orontii. Equations were developed to describe conidial germination with a coefficient of determination (R²) of 0.85 and 0.90 for P. xanthii and G. orontii, respectively. Infection equations resulted in R² of 0.94 and 0.93 for P. xanthii and G. orontii, respectively; and for sporulation, R² of 0.75 and 0.76 for P. xanthii and G. orontii respectively, as a function of temperature. These results can be used to develop models for the risk of cucurbit powdery mildew under field conditions.     

Characterization and pathogenicity of <Emphasis Type="Italic">Bipolaris peregianensis</Emphasis>: the causal organism for leaf spot of hybrid bermudagrass in China

Several golf courses established with hybrid bermudagrass (Cynodon dactylon × C. transvaalensis) were surveyed from April 2011 through April 2015 in Hainan Province, China. The hybrid bermudagrass in these golf courses showed a new leaf spot disease, and a filamentous fungus was consistently recovered from the infected leaves. Based on the morphological characteristics of colony color and appearance, shapes of conidiophores and conidia as well as sequences of internal transcribed spacer regions (ITS) and glyceraldehyde 3-phosphate dehydrogenase (GAPDH), the fungus was identified as Bipolaris peregianensis. The pathogenicity test conducted on healthy hybrid bermudagrass produced leaf spot symptoms one week post inoculation. B. peregianensis mycelia grew in a temperature range of 5 to 35 °C with the optimum temperature being 28 °C.     

Comparative colonisation by virulent versus avirulent <Emphasis Type="Italic">Pyricularia oryzae</Emphasis> on wild <Emphasis Type="Italic">Oryza australiensis</Emphasis>

Pyricularia oryzae (rice blast) conidial development at pre-penetration stage determines success or otherwise of infection inside the rice host plants. Studies on conidial germination and growth on the leaf surface in commercial rice (Oryza sativa) report differently, dependent upon host type and level of blast resistance. Although wild rice (O. australiensis) is known to be an alternative host of blast, the interaction between P. oryzae conidia and wild O. australiensis on its leaf surface has not been previously studied. We found significant (P?<?0.001) differences in conidial development between two blast isolates with different virulence in terms of conidial germination, germ tube growth and appressoria formation on both wild and cultivated rice. Conidial germination at 6 h post-inoculation (hpi) for the virulent isolate was significantly (P?<?0.001) delayed. Germ tubes of the avirulent isolate conidia grew significantly (P?<?0.001) faster and with significantly (P?<?0.001) longer germ tubes than from virulent conidia. Appressoria development for the virulent isolate was significantly (P?<?0.001) faster at its later growth stages of 12 and 18 hpi when approximately 100% of germ tubes formed appressoria. In contrast, formation rate of appressoria for the avirulent isolate was significantly (P?<?0.001) slower and only reached 76% of germ tubes forming appressoria. Appressoria formation on O. australiensis was significantly (P?<?0.001) greater than the formation on O. sativa for both virulent and avirulent P. oryzae at 12 hpi, a clear indication that host type influences the extent of appressoria formation.     

Molecular cloning and virus-induced gene silencing of MiASB in the southern root-knot nematode, Meloidogyne incognita

Root-knot nematodes (Meloidogyne spp.), cause serious damage to agricultural production worldwide. In this study, we designed special primers based on the predicted Mitochondrial ATP synthase b subunit gene (ASB) sequence to clone the same gene in M. incognita (MiASB). The identity between the cloned MiASB and the predicted MiASB was as high as 100 %. Using the tobacco rattle virus (TRV)-mediated virus-induced gene silencing (VIGS) system, we delivered MiASB RNAi triggers to the M. incognita feeding site on tomato seedlings, resulting in significantly fewer galls on the seedlings. Sixty days after inoculation with M. incognita, the number of root galls induced on the MiASB silence-treated seedlings was reduced by 64.3 % compared to that on the control seedlings, and reduced by 64.1 % compared to that on untreated control seedlings. This study revealed the MiASB silencing had a positive effect on the control of root-knot nematodes, and MiASB may be associated with the formation of galls caused by the nematode.     

<Emphasis Type="Italic">Aphelenchoides gorganensis</Emphasis> n. sp. (Nematoda: Aphelenchoididae)<Emphasis Type="Italic">,</Emphasis> a new species from Iran

Phytophthora capsici infection of chili pepper seedlings can cause substantial losses due to damping-off and collar rot diseases. Chemical control is no longer effective due to reported resistance development, on top of the related environmental concerns and the consumer demands for reduced use of fungicides. Biological control is a sustainable option, with several agents having been reported to be effective against this pathogen. This research focused on optimizing the application of strain THSW13 of Trichoderma hamatum and a bacterial isolate BJ10–86 with the objectives of improving chili pepper seed germination, reduce damping-off disease incidence, and improve the growth of the seedlings. Bacterial isolate BJ10–86 was subjected to molecular identification and found to be Pseudomonas aeruginosa. Chili pepper seeds treated with the biocontrol agents, individually or in combination, were seeded into commercial nursery media that had been pre-inoculated with P. capsici zoospores. Over a period of 35 days the chili pepper seed treatments significantly (P = 0.008) reduced the disease incidence of seedlings damping-off. Combined application of T. hamatum and P. aeruginosa was the best biocontrol treatment with an area under disease curve of only 36.61 units compared to 92.87 units for the control treatment. Similar results were observed in vitro where T. hamatum and P. aeruginosa synergistically inhibited P. capsici growth by 73.2 %. The inhibition activity of this treatment was similar to mefenoxam treatment, which implies that it is an effective and sustainable alternative for chili pepper seed treatment. The biocontrol seed treatment had no effect on seed germination and seedling growth.     

Factors affecting <Emphasis Type="Italic">Neofuscicoccum ribis</Emphasis> infection and disease progression in blueberry

Botryosphaeria stem blight is an economically important disease of blueberry worldwide. In this study, factors affecting inoculum production, infection and disease progression of Neofusicoccum spp. in blueberries were investigated. Under laboratory conditions conidia of the main three Neofusicoccum species (N. australe, N. parvum and N. ribis) were released from pycnidia at 15–30 °C and under relative humidities (RHs) of 80–100%, with greatest numbers released by N. parvum. The greatest numbers of oozing pycnidia and conidial release occurred at higher temperatures (25–30 °C) and RHs (92–100%). Inoculation of green shoots with different N. parvum and N. ribis conidial concentrations (50 μL of 5 × 10⁴?5 × 10⁶ conidia/ mL) caused 100% incidence but lesion lengths increased with increasing concentrations. Wound age affected N. ribis lesion development, with lesions only observed for 0–7-day-old wounds in soft green shoots and 0–4-day-old wounds for both hard green shoots and trunks. Colonisation length decreased with increasing wound age. Lesions developed on wounded shoots when plants were exposed to 20 or 25 °C and 90 or 100% RH during the early infection processes; and in non-wounded shoots spot-like lesions were observed although N. ribis colonised the stem tissue. Seasons (summer, autumn and winter) had no effect on susceptibility of wounded plants to N. ribis. External lesions only developed in summer-inoculated plants and colonisation length was lower in winter-inoculated plants. Information on host and environmental factors that affect disease development determined by the study will be used to inform the development of control strategies.     

Diversity in Puccinia triticina detected on wheat from 2008 to 2010 and the impact of new races on South African wheat germplasm

Samples of wheat and triticale infected with leaf rust were collected from 2008 to 2010 in South Africa to identify Puccinia triticina races. Races were identified based on their virulence profile on standard differential lines. Eight races were identified from 362 isolates. The dominant races were 3SA133 (syn. PDRS) in 2008 (78 %) and 2009 (34 %), and 3SA145 (47 %) in 2010. Race 3SA145 (CCPS) identified in 2009 was a new race in South Africa with virulence for the adult plant resistance gene Lr37. Another new race, 3SA146 (MCDS), was identified in 2010. Race 3SA146 is also virulent for Lr37 but unlike 3SA145, it is virulent for Lr1 and Lr23 and avirulent for Lr3ka and Lr30. Microsatellite analysis showed that 3SA145 and 3SA146 shared 70 % genetic similarity with each other, but only 30 % similarity with other races in South Africa, suggesting that both represent foreign introductions. In seedling tests of 98 South African winter and spring cultivars and advanced breeding lines, 27 % were susceptible to 3SA145 and 3SA146 but resistant to 3SA133. In greenhouse studies of 59 spring wheat adult plants, 19 % of breeding lines and 46 % of cultivars were susceptible to 3SA145, whereas 29 % of the lines and 53 % of cultivars were susceptible to 3SA146. The cssfr6 gene-specific DNA marker confirmed the presence of Lr34 gene for leaf rust resistance in a homozygous condition in 28 wheat entries. Five entries were heterogeneous for Lr34. Several entries which were susceptible as seedlings to the new races carried Lr34. These lines are expected to show lower levels of leaf rust as adult plants. Results of these studies indicate a continued vulnerability of South African wheat cultivars to new races and emphasise the importance of regular rust monitoring and the need to incorporate genes for durable resistance.     

Effect of Formulation and Application Method on the Efficacy of Aerial and Submerged Conidia of Metarhizium flavoviride for Locust and Grasshopper Control

A study was carried out to investigate the relative infectivity of aerial and submerged conidia of Metarhizium flavoviride to Schistocerca gregaria and Zonocerus variegatus. The effect of formulation and application method on initial infectivity and field persistence of these conidia was investigated. Strain IMI 330189 was highly virulent to S. gregaria but showed relatively low virulence to Z. variegatus. Direct contact with conidia from the initial spray application resulted in 100% mortality of S. gregaria for all formulation and application combinations. The mean survival time of infected locusts was significantly shorter for treatments using a knapsack sprayer containing submerged conidia in water plus 10 ml litre⁻¹ ‘Codacide’™ (seven days), than treatments with aerial conidia in oil using ULV techniques (8.9 days) or submerged conidia in modified (water plus adjuvants) ULV (MULV) (nine days) or in water-based (VLV) applications (9·3 days). Both aerial and submerged conidia persisted long enough in the environment to effect significant mortality via secondary pick-up of spray residue from vegeta-tion. Persistence was greatest in the ULV and MULV treatments, where the oil component of the formulations provided greater protection of the conidia from environmental stresses. The consequences of secondary pick-up of conidia from the different treatments on total mortality from a single application were examined using a simple host–pathogen model. This predicted that the ULV treatment would be much more effective than the other treatments under conditions where direct contact with the spray was limited. The results of these investigations are discussed in the context of development of optimum spray strategies for control of locusts and grasshoppers, and other pests, under different environmental conditions.     

Potential distribution of citrus black spot in the United States based on climatic conditions

Citrus black spot (CBS), caused by Phyllosticta citricarpa McAlp Van der Aa, was recently detected in southern Florida in the US. In addition to infected plant propagation materials, movement of infected citrus fruit poses a concern for potential spread of the disease out of the current quarantine zone, because lesions with pycnidia and conidia could develop after harvest. The conditions conducive for mycelial growth and development of pycnidia and conidia are not well known. Therefore, effects of temperature and relative humidity on growth and conidial production of P. citricarpa were determined and used as parameter inputs in CLIMEX to predict potential establishment of CBS in North America. Colony growth and conidial production in vitro were optimal at 27 °C, whereas there was no growth below 4 °C and above 37 °C. On fruit, lesion development and conidial production were observed at 4 °C, though at a low rate, indicating a greater versatility of the fungus on fruit. More full pycnidia were produced on the CBS lesions at 91 % RH compared to 84 %. Input parameters for CBS risk in CLIMEX obtained from literature, which reflected conditions for infection in spring/summer in Florida, predicted potential establishment in Florida but not in California. However, altering the parameter values to account for survival of the pathogen in leaf litter in winter predicted potential establishment in California as well as Florida. Thus, P. citricarpa could possibly establish beyond Florida if this organism is transported outside of the current quarantine zone to other citrus production areas.     

TaqMan PCR assay for detection and quantification of <Emphasis Type="Italic">Stagonosporopsis tanaceti</Emphasis> in pyrethrum seed and seedlings

Pyrethrum seed has an important role in the transmission of Stagonosporopsis tanaceti, the cause of ray blight disease of pyrethrum. A TaqMan probe based polymerase chain reaction (PCR) assay was developed to quantify the level of S. tanaceti inocula in pyrethrum seed and seedlings. Primer pair (St_qF3, St_qR2) was designed based on the intergenic spacer (IGS) region of S. tanaceti, which produced a 125 bp amplicon specific to S. tanaceti. TaqMan PCR assay using St_qF3, St_qR2 and a probe St_qP was highly specific against the genomic DNA of S. tanaceti, but did not amplify DNA of 14 related Stagonosporopsis species or other foliar pathogens of pyrethrum. The sensitivity limit of this assay was measured using the cycle threshold (C_t) value which ranged from 17.59 for 10 nanograms (ng) to 36.34 for 100 femtograms (fg) genomic DNA of S. tanaceti. There was a significant negative correlation (r = ?0.999, P < 0.001) between the C_t value and the percent of S. tanaceti infected seed. In addition, this TaqMan PCR assay detected latent infection within seedlings. This assay could be applied to test commercial seed and seedlings before deciding on the appropriate management practices.     

Interaction Between Entomopathogenic Fungi and Some Aphid Species in Thailand

Seven aphid species were screened with twelve entomopathogenic fungi for the selection of the most effective fungus. Beauveria bassiana CKB-048 caused the highest mortalities for all aphid species tested in their nymphal stage. The cowpea aphid (Aphis craccivora Koch) was chosen for further studies in its adult stage using B. bassiana CKB-048 as a biocontrol agent, and carbosulfan was used for a comparison in greenhouse testing. The mode of action of the fungal killing was studied by scanning electron microscopy. The endophytic colonization of the cowpea plant by B. bassiana CKB-048 was studied and confirmed using both stereo microscopy and transmission electron microscopy. The chitinase enzyme activity of cowpea plants after they were sprayed with the fungus or carbosulfan was found to fluctuate throughout five days. The lethal concentrations of B. bassiana CKB-048 suspensions that killed 50 % (LC₅₀) of A. craccivora was 6.69 × 10⁷ conidia/ml for the nymphs and 8.25 × 10⁷ conidia/ml for the adults. The time for lethality (LT₅₀) with a concentration of 2 × 10⁸ conidia/ml was 3.25 days for nymphs and 4 days for adults.