The resistance of plants to powdery mildew.
 
The question, why a plant of one species can be attacked by a parasite and an other plant not, could not be answered until now, for science has no explanation found yet. At the University of Gießen a new resistance mechanism was found by Dr. Karl-Heinz Temmen after several years of investigation of the resistance of plants against powdery mildew (TEMMEN 1977, TEMMEN et al 1979c).

Powdery mildew is one of the most important obligate prasites in plant production which attacks leaves, flowers, young stems of mostly all cultivars and causes death of growing points, distortion of leaves and the stunting of shoots. Based on susceptible and resistant plant species, cultivars and clones, the resistance mechanism to powdery mildew was investigated on leaves of the following cultivars: black currant (Ribes) apple (Malus), rose(Rosa), cucumber(Cucumis) and also barley (Hordeum). It was discovered, that the youngest leaves in the unfolding stage of all dicotyles are always susceptible. With further development, tolerance to the fungus increases, but to a different degree in variety and clones of cultivars. On some plants, leaves 1 to 3, numbered basipetally, are susceptible and have macroscopically visible fungus colonies, while on other plants susceptibility extents to leaves 18 – 20. The histological evaluation of all cultivars , based on safranin-stained sections, revealed that susceptible leaves had a high percentage of cells completely filled with cytoplasm, compared to epidermal cells of resistant leaf stages, which were either partially or completely vacuolized. When the percentage of these differently stained cells was determined, a satisfactory correlation between stainability and susceptibility became apparent. The cell development of epidermal cells on monocotyledons comparing to dicotyledons is different because the ontogenetic development of the embryonal epidermal cells in monocotyledons starts within the stalkknots . The stomata cells, the auxiliary cells and the short cells develop from the jung epidermal cells. Each stomata cell with two auxiliary cells is surrounded by 4 short cells which are in neighbourhood of long cells. After development of the cells the vacuolization is almost finished exept in in some stomata and short cells which we call “starter cells”. Those cells are completely or at least partially filled with cytoplasm and are essential to initiate fungal development. To be able to draw their nutrients even from the vacuolized epidermal cells fingerlike prolongations of the haustoria are being formed.

How can it be interpreted?
The natural senescence of cells has long been known and is a matter for botanical text-books. But the phenomenon of the difference in susceptibility in correlation to the degree of vaculozation of epidermal cells of suscetible and resistant cultivars has been observed for the first time by Dr. Karl-Heinz Temmen.
This means the powdery mildew fungus depends on a sufficient supply of nutrients in epidermal cells. So the formation of primary haustoria require epidermal cells rich in cytoplasm. These are called “starter cells”, which correspond to the auxiliary cells of barley cultivars. A very early contact of primary haustoria with cytoplsm is necessary for further fungal development. When secondary mycelium has been formed, the fungus attempts to establish haustoria also in neighbouring cells, in order to secure the nutritional basis. Such secondary haustoria are formed even in completely vacuolized cells, which do not allow formation of primary haustoria. Such relation has been observed in all host-parsite-combinations mentioned before. This “lack of cytoplasm-effect” is strengthened by the fact, that the fungus can develop on epidermal cells above leaf veins of leaves which do not permit powdery mildew on interveinal areas. Epidermal cells above leaf veins are mostly completely filled with cytoplasm, while the neighbouring cells above interveinal areas are vacuolized.

How does this work?
A great number of Ribes cv, Malus cv, Rosa cv , Cumumis cv and Hordeum cv were tested for their susceptibility. The differences in susceptibility in all models are linked in the degree of vacuolisation of epidermal cells. The more pronounced the vacuolization, and the greater the quantity of the host cytoplasm, the better the fungus thrives. This can be seen from the fact that powdery mildew generally thrives better on leaves of well-nourished plants than on leaves of specially N defiency. For demonstration matter the susceptible cultivar “Super Star” and the resistent cultivar “Gloria Dei” were choosen. No macroscopically visible mildew colonies were detectable on the resistant cv, while the susceptible cv was heavily infected up to pinnate leaf No. 9, when numbered basipetally (Fig. 1). Although mildew colonies were macroscopically invisible on some leaf stages fungal development could be detected by microscopic examination using the Technicoll technique, which is based on adherence of fungal structure to the adhesive. On leaf No. 5 of the resistent cultivar “Gloria Dei” with cells mostly in the stretching stage the germination, appressoria formation and appearance of secondary hyphae of the fungus occur within approximately the same time range as with the succeptable cultivar “Super Star”. However in opposite to “Super Star” on “Gloria Dei” only conidiophores are formed but no conidia are produced. A few hours later hyphae and coniophores shrivel and become dry (Fig. 2). The histological evaluation, based on safranin-stained sections, revealed that susceptible leaves had a high percentage of epidermal cells complelely filled with cytoplasm, compard to epidermal cells of resistant leaf stages, which were either partially or comletely vacuolized. Top leaflets of the first pinnate leaf of susceptible cv had more than 50 % non-vacuolized cells, whereas corresponding leaves of resistant cv had 30 %. In leaves No 3, 5, 7, 9 vacuolization of epidermal cells steadily progessed; the depletion of cytoplasm however occured at a faster rate in cells of resistant cv (Fig. 3).
What do these differences in cytoplasm content mean?
The degree of vacuolization or depletion of cytoplasm has an influence on fungal development In the cells packed with cytoplasm the fungus penetrated and formed haustoria 24 hrs p.i., however papillae were rarely observed. In cells almost completely vacuolized, the fungus had formed only penetration pegs 24 hrs p.i., which had become surrounded by papillae. After prolonged incubation, the number of normal haustoria had increased in vacuolized cells 144 hrs p.i., penetration of papillae, however, was not observed.
It appears, that the formation of primary haustoria require epidermal cells rich in cytoplasm (starter cells), which would correspondent to the auxiliary cells of Hordeum cv considered essential for the primary infection by HIRATA and TOGASHI (1957). A low transport of nutrients in the “starter cells” take place during the development of bodies of primary haustoria and a high rate of transfer of substances coincides with the time when bodies have attained full size (MOUNT and ELLINGBOE 1969).That would explain the question why a very early contact of the primary haustorium with cytoplasm is necessary for further fungal attempts to establish haustoria also in neighbouring cells, in order to secure the nutritional basis. Apparently one haustorium is not sufficient to allow conidia production (HIRATA; 1967, 1971). Such secundary haustoria are quite readily formed even in completely vacuolized cells, which previously did not allow formation of primary haustoria. Such a relation has been observed in all four host-parasite-combinations mentioned before. A “starter cell”, rich in cytoplasm, appears to be a prerequisite for initial fungal development. Once a primary infection has been established, the fungus can make use of more or less vacuolized neighbouring cells by secundary haustoria. Therefore resistant leaves have less epidermal cells with high cytoplasm content, necessary for the establishment of the primary infection. Chances to encounter a “starter cell” are thus considerably reduced. With leaf development the number of “starter cells” decreases further, thus making the leaves even more inhospitable for the fungus. Fungal development is not completely suppressed, but retarded and reduced to such an extent that it appears to nullify its epidemiological importance. Likewise, there is little or no damage on the attacked leaves. This condition is considered the basis for horizontal reistance of plants to powdery mildew.

What is the meaning of the frequently observed papillae?
These structures are formed in response to invading pathogens in cells of susceptible and resistant host plants (HEATH 1974), as well as in cells of non-host plants (RIDE and PEARCE 1979). At the present state of knowledge papillae seem to constitute impregnable mechanical barriers against further fungal penetration. This interpretation is consistent with the results of most histological investigations, but a question remains unanswered: why, e.g., can primary infection not develop haustoria in vacuolized cells but only papillae, whereas secundary infections can do so readily? Even though these questions cannot be answered at the moment it seems that vacuolized cells cannot deliver nutrients for the development of a primary haustorium so that the hostcell is able to repair the damage during penetration by forming a papillae.

What is the meaning of horzontal and vertical resistance?
There are 3 host-parasite-interactions to be differentiated:
  1. vertical - or race specific resistance is predestinated not for haustoria formation but only for papillae (Fig. 4a). This means, that either a fungal race with sufficient virulence does not exist or it is a question of a non-host-interaction. For example the rose cv “Super Star”, first introduced in Great Britain, was resistant to powdery mildew for about 10 years, when it suddenly became susceptible. This example is not an argument against the postulated horizontal resistance. According to the percentage of epidermal cells rich in cytoplasm this cv has to be considered susceptible. Due to absence of a fungal race with sufficient virulence, this cv was resistant during the early years of cultivation. Thus it constitutes an example for vertical resistance which must not be confused with the horizontal resistance exhibited by “Gloria Dei”.
  2. Horizontal resistance is predestinated by primary haustoria in cells rich in cytoplasm (starter cells), formation of papillae during primary infection in cells poor of cytoplasm, forming of secundary haustoria in all kind of epidermal cells, absence of sporulation because of few cells rich in cytoplasm. (Fig. 4b).
  3. Susceptibility occurs on those leaves of which cells are rich in cytoplasm. Primary haustoria find sufficient cells with high cytoplasm content. Papillae are rare and sporulation ist normal because of enough cells rich in cytoplasm (Fig. 4c).
 
References:
  1. TEMMEN, Karl-Heinz, 1977
    Investigation on Ribes cv against powdery mildew (Dissertation)
  2. TEMMEN, K.-H., GRUPPE, W. and SCHLÖSSER, E. (1979c)
    Investigations on the resistance of plants to powdery mildew. III. The basis for
    horizontal reistance of Ribes spp.
    in Plant Disease (1980) How Plants Defend Themselves, p.164 Academic Press,
    New York- London, Toronto, Sydney, San Francisco, 1980
  3. HIRATA, K. & TOGASHI, K., 1957
    in R.M. SPENCER (ed.): the Powdery Mildews, p. 268
    Academic Press, London-New York-San Francisco, 1978
  4. MOUNT S.M. & ELLINGBOE, A.H.
    Phytopahtology, 1969, 59, 235
  5. HIRATA, K., 1967, 1971
    in R.M. SPENCER (ed.). The Powdery Mildews, p. 204
  6. HEATH. M.
    Physiol. Plant Path., 1974, 4 , 303-414
  7. RIDE, J.P. & PEARCE. R.B.
    Physiol. Pl. Path. 1979,15, 79-92