Use of the temporary immersion bioreactor system (RITA<Superscript>®</Superscript>) for production of commercial <Emphasis Type="Italic">Eucalyptus</Emphasis> clones in Mondi Forests (SA)

In order to optimise tissue culture systems and to meet production targets, Mondi Forests biotechnology programme has in the last 2years concentrated efforts on the use of the RITA^® temporary immersion bioreactor system. Protocols have been established for six Eucalyptus clones. Results indicate a four- to six-fold increase in yield, in half the time, with the RITA^® system when compared with axillary bud proliferation on semi-solid media. Furthermore, plants produced from the RITA^® system are hardier and acclimatize better, giving higher yields of hardened-off plants. The establishment of aseptic axillary shoots into the RITA^® system is from shoots in the semi-solid system. Highest multiplication was achieved using 30-second flushes of medium every 10min, starting with 50 shoots per vessel. The multiplication cycles in RITA^® are between 14 and 18days, compared with 25–28days in a semi-solid system. There is minimal callus evident on the leaves and bases of the stems of plants in the RITA^® system and, in addition, cold-tolerant plants have a greater rooting competence when compared with plants coming from the semi-solid system. Ex vitro rooting of RITA^® – derived plantlets is substantially better than the plants from the semi-solid media.     

In vitro propagation of three <Emphasis Type="Italic">Agave</Emphasis> species used for liquor distillation and three for landscape

In vitro bud clusters of Calathea orbifolia (Linden) Kennedy were obtained and subcultured in semi-solid (agar) medium and temporary immersion system (TIS) for 12 weeks. Uniform young plants were selected and transferred to soilless mix in a growth chamber for ex vitro acclimatization during 35 days, followed by growing in a shaded greenhouse for 65 days. Comparison of in vitro leaf anatomy, ex vitro photosynthetic behaviors and growth was made between two cultural systems. Plants in TIS produced thicker leaf chlorenchyma and aquiferous parenchyma, lower stomatal frequency and more epicuticular wax than did those in semi-solid medium. Plants from semi-solid medium had consistently lower leaf Fv/Fm values than plants from TIS. Leaf Fv/Fm value in plants from TIS decreased to 0.65 at day 7 after transfer and increased soon up to 0.76 thereafter. In contrast, leaf Fv/Fm value in plants from semi-solid medium reduced to 0.27 at day 7 after transfer and increased slowly up to 0.68 at day 35. During ex vitro acclimatization, plants in TIS had substantial higher photosynthetic rates than plants in semi-solid medium. Plants from TIS had subsequent higher leaf area, fresh and dry weights than plants from semi-solid medium.     

Temporary immersion systems in plant micropropagation

Temporary immersion systems for plant micropropagation have been described and grouped into 4 categories according to operation: tilting and rocker machines; complete immersion of plant material and renewal of the nutrient medium; partial immersion and a liquid nutrient renewal mechanism; complete immersion by pneumatic driven transfer of liquid medium and without nutrient medium renewal. The positive effects of temporary immersion on micropropagation are indicated for shoot proliferation and microcuttings, microtuberization and somatic embryogenesis. Immersion time, i.e. duration or frequency, is the most decisive parameter for system efficiency. Optimizing the volume of nutrient medium and the volume of the culture container also substantially improves efficacy, especially for shoot proliferation. Temporary immersion also generally improves plant material quality. It results in increased shoot vigour and in the frequency of morphologically normal somatic embryos. Hyperhydricity, which seriously affects cultures in liquid medium, can be eliminated with these culture systems or controlled by adjusting the immersion times. Plant material propagated by temporary immersion can perform better during the acclimatization phase than material obtained on semi-solid or in liquid media. Successful regeneration of plants, after direct sowing on soil of Solanum tuberosum microtubers and Coffea arabica somatic embryos produced in temporary immersion bioreactors, has been demonstrated. As could be expected when using liquid medium for micropropagation, several estimations confirm large gains in efficacy from temporary immersion. The parameters most involved in reducing production costs include: (1) the drastic reduction in work; (2) reduction in shelving area; (3) reduction in the number of containers used; (4) better biological yields. Scaling-up somatic embryogenesis and shoot proliferation procedures involving temporary immersion systems in order to commercialize this process are now taking place.     

Mineral nutrient supply, cell wall adjustment and the control of leaf growth

The possibility that changes in the plasticity of expanding cell walls are involved in regulating early leaf growth responses to nutrient deficiencies in monocot plants was investigated. Intact maize seedlings (Zea mays L.) which were hydroponically grown with their roots in low-nutrient solution (1 mol m^?3 CaCl₂) showed early inhibition of first-leaf growth, as compared with seedlings on complete nutrient solution. This early inhibition of leaf growth was not associated with reduced cell production. However, segmental elongation along the cell expansion zone at the base of the leaf and the lengths of mature epidermal cells were reduced by the low-nutrient treatment. Solute (osmotic) potentials in the expanding leaf tissues were unchanged. In contrast, low-nutrient treatments significantly altered leaf plasticity, i.e. the irreversible extension caused by applying a small force in the direction of leaf growth. For example, in vivo plasticity decreased, along with leaf growth, after transfer of seedlings from complete nutrient solution to low-nutrient solution for 15 h. Conversely, in vivo plasticity increased, along with leaf growth, after transfer of plants previously grown on low-nutrient solution to complete nutrient solution for 15 h. The nutrient treatments also induced similar changes in the in vitro plasticity of the expanding leaf cell walls. There were no consistent changes in elasticity. Thus, reductions in the plasticity of expanding leaf cell walls appear to be involved in controlling the early inhibition of maize leaf growth by root imposition of nutrient stress.     

Cryopreservation of encapsulated gentian axillary buds following 2 step-preculture with sucrose and desiccation

Alginate beads containing axillary buds of in vitro-grown gentian (Gentiana scabra Bunge var. buergeri Maxim.), were successfully cryopreserved following 2 step-preculture with sucrose and desiccation. The optimal preculture conditions were as follows: axillary buds were excised from in vitro-grown gentian plants and precultured on semi-solid Murashige and Skoog (MS) medium containing 0.1 M sucrose for 10 days (25 °C, 16-h photoperiod) (first step). This was followed by incubation on semi-solid MS media containing 0.4 M (1 day) and then 0.7 M sucrose (1 day) (second step). After preculture, the buds were encapsulated in alginate beads and desiccated aseptically on silica gel for 9 h to a water content of 10% (fresh weight basis), followed by immersion in liquid nitrogen (LN). With this protocol, 87% of the gentian buds survived exposure to LN and showed normal development of shoots and roots in vitro and in vivo. Depletion of NH₄NO₃ in the regeneration medium did not improve survival following desiccation and exposure to LN. The results show that 2 step-preculture with sucrose is effectively applicable in encapsulation–desiccation based cryopreservation of gentian axillary buds. This preculture can replace the conventionally used lengthy cold-hardening treatment and is useful for routine cryopreservation of gentian germplasm.     

Improving micropropagation of Mentha × piperita L. using a liquid culture system

In the current study, in vitro shoot proliferation and plant regeneration of Mentha × piperita L. (peppermint) cultivar ‘Black Mitcham’ was compared in semi-solid and liquid culture systems. Shoot tips from field-grown plants were used as explants to study shoot proliferation response on either Murashige and Skoog (MS) or Chee and Pool (C2D) medium containing varying levels of 6-benzylaminopurine (BAP), kinetin, and 6-γ,γ-dimethylallyl aminopurine (2iP). Differences in leaf ultrastructure and antioxidant capacity of greenhouse-grown and micropropagation-derived plants were studied to identify potential changes occurring during in vitro culture. Among the various media treatments tested, the maximum number of shoots was produced on the C2D medium with 4.0 μM BAP (40.7) followed by the MS medium with 4.0 μM BAP (32.2). Among the rooting treatments, shoots on the MS medium with 1.0 μM indole-3-butyric acid (IBA) produced the maximum number of roots (14.4). The number of shoots produced in Liquid Lab Rocker® (LLR) vessels containing liquid C2D medium with BAP (103.4) was significantly higher than that produced on semi-solid medium (40.7). No differences were observed in the leaf ultrastructure and antioxidant capacity of leaf extracts obtained from greenhouse-grown and micropropagation-derived plants. The study indicates that the liquid culture system under the described conditions can enhance peppermint micropropagation, with plant material being potentially valuable for use in herbal supplements and essential oil production.

     

Shoot regeneration from leaf explants of five strawberry (<Emphasis Type="Italic">Fragaria × Ananassa</Emphasis>) cultivars in temporary immersion bioreactor system

Summary A temporary immersion bioreactor system (TIB system) provides a convenient and efficient way to propagate plant material in vitro while requiring significantly lower labor input than conventional methods. The applicability of a TIB system for adventitious shoot regeneration from strawberry leaf explants was studied. Five commercial cultivars, i.e. Bounty, Jonsok, Korona, Polka, and Zephyr, were propagated in regeneration medium in commercially available TIB bioreactors (RITA^?) and, for comparison, on the same medium solidified with agar. The TIB system proved to be well suited for shoot propagation and for subsequent subculture of the developing plantlets. Regeneration frequencies were 70±8 to 94±2% and 83±5 to 92±3% in the TIB system and on semi-solid medium, respectively. The labor time taken by the TIB system was less than half of the time required for handling plant material for cultivation on semi-solid medium. This system thus provides a convenient method that could be adopted for commercial in vitro propagation or for regeneration of transgenic strawberry cultivars.     

Phenotypic responses of <Emphasis Type="Italic">Spartina anglica</Emphasis> to duration of tidal immersion

Although Spartina anglica C.E. Hubbard continues to be invasive in many countries, this species has experienced a drastic decline in coastal China over the last decade. We hypothesize that changes in the duration of tidal immersion were responsible for this decline because the elevation of the S. anglica-dominated area in coastal China has increased greatly over the last decade. We examined the effects of the duration of simulated tidal immersion and plant material provenance on growth, asexual reproduction, biomass accumulation, and allocation (percent of above-ground biomass to total biomass) of S. anglica in a greenhouse experiment. The provenance of S. anglica did not significantly affect any traits measured except for height, stalk diameter, and leaf area. However, all traits were affected by the duration of immersion. Plants grown under 6 h of immersion were taller and had more leaves, more roots, and larger leaf area than those under 2, 4, 8, and 10 h of immersion. Asexual traits and biomass of the plants grown under 6 h of immersion were significantly larger than those under other immersion durations. The results suggested that S. anglica benefits from tidal immersion and decreasing duration of tidal immersion may have resulted in the decline of the S. anglica populations in coastal China. Thus, controlling the duration of tidal immersion may be an effective way of controlling invasiveness of this species elsewhere in the world.     

Protoplast isolation, culture, and fusion protocols for kenaf (Hibiscus cannabinus)

Protoplast isolation and culture protocols were developed for ten cultivars of Hibiscus cannabinus L. (kenaf). Leaves from seedling lines maintained in vitro were used as donor tissues. Optimal cell wall digestions were achieved with a combination of cellulysin (1.0%) and macerase (0.5%). Average yields ranged from 0.9×10⁵ to 5.9×10⁶ protoplasts g fw^-1 leaf tissue with viability estimates ranging from 53% to 87%. This protocol was ineffective for leaf tissue taken from plants grown in vivo. Protoplasts harvested from plantlets maintained in vitro produced rapidly growing calluses when plated in semi-solid medium after an initial culture in liquid medium. First cell divisions were observed within four to six days after initial culture in medium containing plant growth regulators 2,4-dichlorophenoxyacetic acid (1.4 M) and kinetin (13.8 M). An electrofusion protocol which did not significantly reduce protoplast viabilities was developed for kenaf protoplasts. The maximum fusion frequency (4.6%) was obtained with an electrofusion voltage of 2.0 kV cm^-1.Abbreviations 2,4-d 2,4-dichlorophenoxyacetic acid - BA 6-benzylaminopurine - FDA fluorescein diacetate - MS Murashige and Skoog - NAA 1-naphthaleneacetic acid - PGRs plant growth regulators - SCL seed clonal line     

A Comparison of the Disease Reactions of Stems and Detached Leaves of Soil and in vitro Grown Plants and Regenerants of Oilseed Rape to Leptosphaeria maculans and Protocols for Selection for Novel Disease Resistance

Protocols for selecting plant tissues of winter oilseed rape (Brassica napus subsp. oleifera) with resistance to Leptosphaeria maculans by either stem or leaf inoculation of both soil and in vitro grown plant material are described. The stem inoculation procedure gave good correlation (r = 0. 92) between the 50 day stem disease scores of eight out of nine cultivars of soil grown winter oilseed rape inoculated with isolate 41A4 of L. maculans and the N. A. B. esistance ratings or resistance data from field trials. The exception was the cultivar Liradonna. Inoculation of stems of five cultivars with isolates 41A4, 433 and 478 indicated a range of isolate virulence 478 > 41A4 > 433. This was the inverse of that observed in leaf inoculations. Application of the stem inoculation procedure to in vitro shoot cultures allowed differentiation of resistant and susceptible cultivars, including the cultivar Liradonna, after 20 days incubation at 20°C. The protocol was also applicable to plantlets regenerated from thin cell layer explants grown in vitro. Inoculations with isolate 433 allowed the differentiation of resistant, intermediately resistant and susceptible leaf material of soil grown plants, when leaf discs from young leaves were incubated on water agar supplemented with BAP (1 × 10^?5 M) at 25°C for 10 days. Intermediately resistant leaves were resistant after 10 days and susceptible after 15 days of incubation. Leaves of shoot cultures grown in vitro were more susceptible than the corresponding soil grown material. However, inoculation of old leaves with isolate 41A4 (an isolate of less virulence on leaves than 433) distinguished the cultivars after 15 days of incubation. These protocols allow the accurate assessment of resistance to L. maculans at the stem or leaf level and are of use in traditional as well as in vitro selection programmes.     

Comparative photosynthesis,growth, productivity,and nutrient use efficiency among tall- and short-stemmed rain-fed cassava cultivars

Field trials under rain-fed conditions at the International Center for Tropical Agriculture (CIAT) in Colombia were conducted to study the comparative leaf photosynthesis, growth, yield, and nutrient use efficiency in two groups of cassava cultivars representing tall (large leaf canopy and shoot biomass) and short (small leaf canopy and shoot biomass) plant types. Using the standard plant density (10,000 plants ha⁻¹), tall cultivars produced higher shoot biomass, larger seasonal leaf area indices (LAIs) and greater final storage root yields than the short cultivars. At six months after planting, yields were similar in both plant types with the short ones tending to form and fill storage roots at a much earlier time in their growth stage. Root yield, shoot and total biomass in all cultivars were significantly correlated with seasonal average LAI. Short cultivars maintained lower than optimal LAI for yield. Seasonal P _N, across cultivars, was 12% greater in short types, with maximum values obtained in Brazilian genotypes. This difference in P _N was attributed to nonstomatal factors (i.e., anatomical/biochemical mesophyll characteristics). Compared with tall cultivars, short ones had 14 to 24 % greater nutrient use efficiency (NUE) in terms of storage root production. The lesser NUE in tall plants was attributed mainly to more total nutrient uptake than in short cultivars. It was concluded that short-stemmed cultivars are superior in producing dry matter in their storage roots per unit nutrient absorbed, making them advantageous for soil fertility conservation while their yields approach those in tall types. It was recommended that breeding programs should focus on selection for more efficient short- to medium-stemmed genotypes since resource-limited cassava farmers rarely apply agrochemicals nor recycle residual parts of the crop back to the soil. Such improved short types were expected to surpass tall types in yields when grown at higher than standard plant population densities (>10,000 plants ha⁻¹) in order to maximize irradiance interception. Below a certain population density (<10,000 plants ha⁻¹), tall cultivars should be planted. Findings were discussed in relation to cultivation and cropping systems strategies for water and nutrient conservation and use efficiencies under stressful environments as well as under predicted water deficits in the tropics caused by trends in global climate change. Cassava is expected to play a major role in food and biofuel production due to its high photosynthetic capacity and its ability to conserve water as compared to major cereal grain crops. The interdisciplinary/interinstitutions research reported here, including an associated release of a drought-tolerant, short-stem cultivar that was eagerly accepted by cassava farmers, reflects well on the productivity of the CIAT international research in Cali, Colombia.     

Acclimation of tobacco plantlets to ex vitro conditions as affected by application of abscisic acid

Plantlets of Nicotiana tabacum L. cv. Petit Havana SR1 were grown in vitro on Murashige and Skoog medium containing 2% saccharose, and then transplanted ex vitro into pots with coarse sand and Hewitt nutrient solution. In the first day after transplantation, the anti-transpirant abscisic acid (ABA; 0.01, 0.05 or 0.10 mM) was added to the substrate. Leaf stomatal conductance (gs), which was high in plants during the first days after transplantation similarly as in plantlets grown in vitro, was considerably decreased by ABA-treatment. However, in the further days gs decreased more quickly in control than in ABA-treated plants, and after 2 or 3 weeks gs was significantly lower than that of plantlets grown in vitro but similar in control and ABA-treated plants. Two weeks after transplantation, net photosynthetic rate, chlorophyll a + b content, maximal photochemical efficiency, and actual quantum yield of photosystem II in plant leaves were higher in comparison with those in plantlets grown in vitro. ABA-treatment had slight positive or insignificant effect on photosynthetic parameters and enhanced plant growth. Thus ABA application can alleviate 'transplant shock' and speed up acclimation of plantlets to ex vitro conditions.     

Effect of nodulation on the nitrate assimilation in vegetative soybean plants

Summary Nitrate assimilation in the first trifoliate leaf of vegetative soybean plants (Glycine max L. Merr, cv Hodgson) was studied in relation to nodulation. Nodulated and non-nodulated plants were grown in a nitrate medium (4 mM). As a control nodulated plants were grown in a nutrient medium without combined nitrogen. This study included measurements of the acetylene reduction activity of the whole plant and of thein vitro nitrate reductase, glutamine synthetase and glutamate dehydrogenase activities in the first leaf and of the nitrate concentration. Nitrate accumulation and nitrate reductase activity were depressed in nodulated plants; root growth was decreased in the presence of nitrate. The relationships between nitrate assimilation and nodulation are discussed.     

Development of a temporary immersion system (RITA®) for mass production of sugarcane (<Emphasis Type="Italic">Saccharum</Emphasis> spp. interspecific hybrids)

Commercial micropropagation of sugarcane is largely determined by the clonal fidelity and the cost of plants produced. Rapid production of plants in vitro reduces the frequency of offtypes in many species. By exploiting the concept of transverse thin cell layer culture, we have developed a rapid, high frequency direct plant regeneration system, called SmartSett®, for commercial sugarcane cultivars grown in Australia. Similar to conventional micropropagation, labour remains the major cost of this plant production system. Hence, to reduce the labour component, we have integrated the SmartSett® system with the RITA® temporary immersion bioreactor. Thin transverse leaf sections or fragmented leaves cultured on agar-based SmartSett® shoot induction medium were used as the starting material for RITA®. Shoot initiation on semi-solid medium prior to transferring to RITA®, culture immersion frequency, explant size and genotype determined the productivity (number of plants produced per unit culture) of the system. Results obtained with cultivar Q165 indicate that explants cultured for 45 d on SmartSett® shoot induction medium were the most prolific, producing on average 275 shoots per vessel after 45 d of culture in RITA with 1 min immersion every 12 or 24 h. Using the fragmented tissue, 14-d-old explants and 3-mm leaf tissue fragments were the most productive. Experiments with three cultivars (Q117, Q165 and Q205) showed that RITA® culture conditions need to be optimised for each cultivar for maximum plant production.     

Mechanism of resistance to the spotted stalk borer,Chilo sacchariphagus,in the sugarcane cultivar R570

Resistance in sugarcane [Saccharum spec. (Poaceae)] to the spotted stalk borer, Chilo sacchariphagus (Bojer) (Lepidoptera: Pyralidae), was studied by comparing feeding behaviour on resistant cv. R570 and susceptible cv. R579. In a field survey, the feeding behaviour of C. sacchariphagus larvae was described to identify their feeding sites on the plant. In a greenhouse artificial infestation study, we compared the establishment of larvae on potted plants. In laboratory choice and no‐choice experiments, we studied the establishment of larvae on plant organs (stalk, sheath, leaf spindle). Study of the feeding behaviour showed that: (1) first to fourth instars are able to feed on stalk, sheath, and leaf spindle, (2) boring into the stalk occurs mostly in the four uppermost internodes, and (3) most young larvae bore through the abaxial surface of leaf sheaths to reach the stalk. In greenhouse experiments, we observed an early two‐fold reduction of the number of larvae on R570 plants within the first 48 h after infestation. In laboratory experiments, larval antixenosis was demonstrated at the abaxial surface of R570 leaf sheath, but was observed neither in the leaf spindle nor in the stalk. First, second, and third instars were susceptible to this antixenosis. We hypothesize that the main resistance mechanism in R570 is an early reduction of larval establishment on plants, due to antixenosis located at the abaxial surface of leaf sheaths.     

Physiological effects of temporary immersion on Hevea brasiliensis callus

In vitro culture by temporary immersion generates potentially stressful conditions for explants that may differ from those associated with classic methods. In order to evaluate the effects of these conditions on physiological changes in explants, different parameters of metabolic activity were investigated for a friable embryogenic callus of Hevea brasiliensis (Müll. Arg.), in response to 1 min, and 1, 12 and 24 h per day of immersion, using semi-solid and agitated liquid media as controls. The relative growth rate of the callus was not significantly different for the 1 min immersion treatment and the controls, but it decreased by about 60% for the 1-, 12- and 24-h immersion treatments. During the immersed stage, the rate of respiration of the callus was comparable for all the treatments. However, during the emersed stage, the respiration rate increased by 140 and 164% for the 12- and 24-h immersion treatments, respectively. Meanwhile, the total adenylate nucleotide concentration and the ratio of ATP/ADP remained almost constant, or even decreased. The adenylate energy charge was comparable for all the treatments, averaging 0.88. The superoxide dismutase activity and the lipid peroxidation increased with the immersion duration, and were significantly higher for the 12- and 24-h immersion treatments than for controls. However, after 24 h in emersed stage, there was no lipid peroxidation, regardless of previous immersion duration. It appears from these results that the immersed stage induced a substantial oxidative stress, which was not associated with the callus immersion per se.     

Lower marine fungi (labyrinthulomycetes) and the decay of mangrove leaf litter

This paper deals with the association of members of the Labyrinthulomycetes (Thraustochytriales and Labyrinthulales) with decaying or decayed leaves at an intertidal mangrove at Morib, Malaysia. Representatives of both orders of these obligately marine unicellular eukaryotes of unresolved taxonomic affinities (Chamberlain & Moss, 1998) were consistently isolated from leaves at all stages of decay from the recently fallen to those in an advanced stage of decay, but not from either green or senescent yellow leaves attached to trees. Baiting experiments using -irradiated leaf discs of Sonneratia and Rhizophora spp. immersed in the aquatic environment of the mangrove, revealed that leaf material was colonised by both labyrinthulids and thraustochytrids within 24 hours of immersion at the test site and these organisms were isolated from the leaf material throughout the 14 day study period. In vitro experiments using axenic cultures of three thraustochytrid genera inoculated onto sterile discs of Sonneratia leaves and incubated for 14 days caused loss of both biomass and structural integrity of the leaf material. Freeze fracture, followed by scanning electron microscopy of leaves inoculated with a thraustochytrid and a strain of Labyrinthula, revealed that penetration of the leaf occured after 4 days and that the thraustochytrid was associated with localised degradation of internal leaf tissues. Cellulase production by an isolate of Schizochytrium aggregatum was detected. The results of all the above investigations are discussed with reference to the role of members of the Labyrinthulomycetes in nutrient cycling in the mangrove.     

Antifungal metabolites (monorden, monocillins I, II, III) from Colletotrichum graminicola, a systemic vascular pathogen of maize

Colletotrichum graminicola is a systemic vascular pathogen that causes anthracnose stalk rot and leaf blight of maize. In the course of an effort to explore the potential presence and roles of C. graminicola metabolites in maize, ethyl acetate extracts of solid substrate fermentations of several C. graminicola isolates from Michigan and Illinois were found to be active against Aspergillus flavus and Fusarium verticillioides, both mycotoxin-producing seed-infecting fungal pathogens. Chemical investigations of the extract of one such isolate (NRRL 47511) led to the isolation of known metabolites monorden (also known as radicicol) and monocillins I–III as major components. Monorden and monocillin I displayed in vitro activity against the stalk- and ear-rot pathogen Stenocarpella maydis while only the most abundant metabolite (monorden) showed activity against foliar pathogens Alternaria alternata, Bipolaris zeicola, and Curvularia lunata. Using LC–HRESITOFMS, monorden was detected in steam-sterilized maize stalks and stalk residues inoculated with C. graminicola but not in the necrotic stalk tissues of wound-inoculated plants grown in an environmental chamber. Monorden and monocillin I can bind and inhibit plant Hsp90, a chaperone of R-proteins. It is hypothesized that monorden and monocillins could support the C. graminicola disease cycle by disrupting maize plant defenses and by excluding other fungi from necrotic tissues and crop residues. This is the first report of natural products from C. graminicola, as well as the production of monorden and monocillins by a pathogen of cereals.     

The effect of headspace renewal in a Temporary Immersion Bioreactor on plantain (Musa AAB) shoot proliferation and quality

The positive effect of ventilation of the culture container on in vitro shoot proliferation and quality was already proven for different species. Hereafter we report on the evolution of the headspace during in vitro culture of plantain in a Temporary Immersion Bioreactor (TIB) on the one hand, and culture on semi-solid medium on the other hand. The CO₂ and C₂H₄ concentration reached a maximum of 12% and 0.45 μl l⁻¹, respectively in the control treatment on semi-solid medium, compared to 5.7% CO₂ and 0.06 μl l⁻¹ C₂H₄ in TIB. The minimal O₂-concentration on semi-solid medium was 15.1%, compared to 19.3% in TIB. The multiplication rate was best in TIB, 6.4 compared to 4.3 in semi-solid conditions, and this was also the case for shoot height (4.3 cm compared to 3.3 cm), and leaf number (2.6 compared to 1.6). Moreover shoots produced on semi-solid medium showed distorted leaves. A typical day-night pattern in CO₂ and O₂ concentration was observed in TIB, as well as on semi-solid medium; this is illustrative for the photosynthetic capacity of the plant material produced in both systems.     

Studies on stomatal function, epicuticular wax and stem-root transition region of polyethylene glycol-treated and nontreatedin vitro grape plantlets

Summary Scanning electron microscopy, light microscopy, and gravimetric analysis was used to evaluate stomatal function, epicuticular wax, and the stem-root transition region of grape (Vitis sp. ‘Valiant’) plantlets grownin vitro, polyethylene glycoltreatedin vitro, and greenhouse-grown plants. Scanning electron microscopic studies of leaf surfaces ofin vitro-grown plants showed widely open stomata as compared to leaf stomata of polyethylene glycol-treatedin vitro-cultured and greenhouse-grown plants. Ultrastructurally, leaf epicuticular wax ofin vitro plants was less dense than in their polyethylene-treated and greenhouse counterparts. Quantitatively,in vitro-grown plants had reduced epicuticular was as compared to polyethylene glycol-treated and greenhouse-grown plants. Light microscopic studies showed no obvious differences in the vascular connections in the stem-root transition region ofin vitro-cultured, polyethylene glycol-treatedin vitro-cultured, and greenhouse-grown plants. It is therefore likely that the rapid wilting and desiccation observed after transplantingin vitro grape plantlets is due to their defective stomatal function and reduced epicuticular wax and may not be due to poor water transport associated with vascular connection.