No abstract

Type I trichomes of tomato leaves (Solanum lycopersicum Mill. cv. Moneymaker), as outgrowths of the plant epidermis, are suitable for monitoring infection processes of powdery mildew species using a high-fidelity digital microscope (DM) without fungal staining. On the trichomes, tomato powdery mildew (Pseudoidium neolycopersici L. Kiss) isolate KTP-03 produced a maximum of four vigorously elongated hyphae per conidium, which stopped growth approximately 12 days after inoculation. Single trichome cells, invaded by fungal hyphae at various fungal infection stages during the 12-day period after the inoculation of single conidia, were cut at the bases and directly collected with small precision scissors (i.e., microscissors) held by the manipulator under a DM. Subsequently, suc-polymerase chain reaction (PCR) (reverse transcription (RT)-PCR followed by nested (N)-PCR) was conducted to explore gene expression in the infected trichome. We selected intron-containing genes from tomatoes and powdery mildew fungi for the detection of constitutive gene transcripts, namely plasma membrane H+-ATPase (LHA2) and β-tubulin 2 (TUB2) genes. In suc-PCR, a single band from spliced mRNAs of both LHA2 and TUB2 genes were detected, suggesting that both genes were successfully transcribed in single KTP-03-infected trichomes. With combined primers for both LHA2 and TUB2 (multiplex RT-PCR/N-PCR), two bands were detected through the amplification of intron-spliced mRNAs of both genes. Therefore, our single-trichome cell PCR amplification method is effective for detecting the expression patterns of genes from both tomato and powdery mildew fungus. Combinations of digital microscopy, microscissors, and multiplex RT-PCR/N-PCR amplification techniques will be useful for simultaneously analysing the molecular interactions between plants and powdery mildew fungi at the level of single tomato leaf trichome cells. Also, this employed technique will be of benefit in other plant species and crops, possessing leaf trichome cells, to elucidate the molecular interactions between plants and pathogens.

Powdery mildew fungi infect plant leaves, reducing the yield of infected melon plants. Therefore, an eco-friendly method of controlling powdery mildew in melon plants needs to be developed. A previous study described how the morphological characteristics of the conidiophores of the melon powdery mildew fungus Podosphaera xanthii Pollacci (designated KMP-6N) grown under greenhouse (natural) conditions and red light-emitting diode (LED) irradiation differed from those grown under growth chamber conditions and blue LED irradiation. In the present study, conidiophores with unconstricted conidia under blue light were collected and inoculated onto host leaves through micromanipulation; the unconstricted conidia germinated and infected the leaves, producing vigorously elongated hyphae. The number of conidia collected, the initial times of conidial release from single colonies, and the number of conidia remaining in chains on conidiophores were examined with electrostatic techniques. Under red light, the number of collected conidia gradually increased with the light irradiation period. The initial conidial release occurred between 2 to 4 h; the number of conidia remaining on the conidiophores gradually decreased and, eventually, the conidiophore lengths became shorter. In contrast, under blue light, few conidia were collected at any given time; the number of conidia on the conidiophores gradually increased and, eventually, the conidiophore lengths became longer. Next, the effects of red and blue light on the spread of powdery mildew infection by placing a KMP-6N-infected melon seedling at the centre of a tray containing healthy melon seedlings were examined. Almost all healthy seedlings caused powdery mildew symptoms at ca. 21 days after red light irradiation, whereas only healthy seedlings near the infected seedlings showed symptoms after blue light irradiation. Thus, the spread of melon powdery mildew infection clearly differed between red and blue light irradiation. This is the first report describing the effects of red and blue light on the spread of P. xanthii infection from a single infected seedling to healthy host seedlings; their results provide insight into the ecological mechanisms of powdery mildew conidial scatter from conidiophores.