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In woody plants, the availability of plant material for in vitro use is usually limited to the short active growth period of the mother plants. To overcome this limitation, an effective protocol is described to break the seasonal effect on in vitro axillary bud sprouting in Leucospermum cordifolium ‘Flame Spike’ and Leucospermum ‘Tango’. Shoots were harvested at four different times of the year, sterilized and cultured in forcing solutions containing a quarter-strength of Murashige and Skoog medium salts (MS, 1962), supplemented with benzyladenine (BA) (0–100 mg L−1) and gibberellic acid (GA3) (0 or 10 mg L−1). After 10 days, multinodal explants were cultured on half-strength MS solid growth regulator-free medium or with GA3. The effect of pre-treatment is clear in the case of L. cordifolium ‘Flame Spike’ and although in L. ‘Tango’ was not evident, its use reduced the time required for in vitro establishment. The use of nutritive solutions with and without growth regulators allowed to increase the in vitro axillary buds sprouting and the percentage of new shoots

The anatomy of field, in vitro and acclimatized shoots (leaves and stems) of two cultivars of Leucospermum (L. cordifolium ‘Flame Spike’ and L. ‘Tango’) was compared using light, scanning and transmission electron microscopy. Field plants showed several scleromorphic anatomical structures related to excess solar radiation such as: cuticle thickness, subepidermal collenchyma and sclerenchyma. Furthermore, a large quantity of phenolic deposits present in the cell lumen of various tissues is also a scleromorphic feature. The special conditions during in vitro culture result in plantlets with abnormal morphology and anatomy. These disorders are associated with the gaseous environment in the culture vessels, low irradiance in the incubation chamber and the addition of sucrose, nutrients and growth regulators to the culture medium. After transfer from in vitro to ex vitro conditions, substantial changes in leaf and stem anatomy were observed, above all in cuticle thickness, epidermal characteristics (stomatal and trichome index, and stomatal and pore size), differentiation of leaf mesophyll, chloroplast structure, and amount and localization of phenolic deposits. These changes allowed the plants to adapt to the new environmental conditions. The study of anatomical features of in vitro shoots facilitated adapting the acclimation protocol to predict which plantlet would survive the critical acclimation stage.

In vitro rooting of two Leucospermum cultivars (L. cordifolium ‘Flame Spike’ and L. ‘Tango’) microcuttings was studied with different indole-3-butyric acid (IBA) treatments: A) including in a solid medium (0, 0.2, 1 and 2 mg L−1), B) by dipping the base of the shoots in 500–2500 mg L−1 for 5 s and C) including in a liquid medium (12.5 and 25 mg L−1) for 24–96 h (induction step). After the treatments B and C, shoots were transferred to an auxin-free root elongation medium containing 20 g L−1 sucrose and solidified with 7 g L−1 agar. The long-term presence of IBA in the culture-medium inhibited root development. The culture of shoots in a liquid medium with higher IBA concentrations and the application of IBA pulses at the base of the microcuttings proved to be suitable treatments to induce rhizogenesis. Culturing microcuttings in a growth-regulator free solid medium after IBA treatments (B and C) triggered root initiation and differentiation. The results show that the microcuttings cultured in a liquid medium with IBA had the highest percentages of rooting. The sequence of anatomical changes during rooting was similar in vitro and ex vitro, the origin of the adventitious root in the vascular cambium. In vitro roots with an organized tissue system emerged from the microcutting stems 6 days after the root induction treatments. The acclimatized plantlets showed a suitable root system. Roots were formed ex vitro from the stem vascular cambium and connections were established between the stem vascular bundles and the roots, allowing root elongation. Some modifications were also observed in the basal rooting zone of stems in L. ‘Tango’.

To obtain high prices Leucospermum cut flowers are required to have long, straight and slender stems. In addition, the time of flowering is also important, with Nov. and Dec. being considered periods of high demand in Europe for Cape Flora products. Several cultivars grown commercially in South Africa are genetically inclined to produce shorter stems, whilst most late flowering cultivars from warm production areas also present a challenge to deliver sufficient stem length. The practice of disbudding to extend the marketing window, especially in warm areas, likewise results in shorter stems the following season. To increase the stem length plant growth regulators were applied to single stems of Leucospermum 'Succession II' and 'Soleil' at Arnelia Farms, Hopefield, South Africa. Seven treatments were applied at five different times following pruning, at two weekly intervals. Shoot growth was monitored, while the duration of the harvest period and inflorescence quality was also recorded. The stem length was found to have increased in stems of all treatments compared to the control, for both cultivars. In 'Succession II' five applications of the 100 mg L-1 benzyladenine (BA) and gibberellic acid (GA3) combination extended the stem length by 200 mm compared to the control. In 'Soleil' five applications of 500 mg L-1 GA3 was responsible for a 200 mm stem length increase. Treated stems differed distinctly from that of the control in that a more upright growth manner was observed. The timing of application of the respective growth regulators is however considered important to obtain the required results. The use of growth regulators has shown commercially potential to increase stem length in Leucospermum.

The proteas market began in the Canary Islands in the eighties and now covers 60 ha between Tenerife, La Palma and Gran Canaria. The production in the islands is specially focused on the gender Leucospermum, being the most important: Lp. `Succession I´, Lp. `Succession II´, Lp. `Tango´, Lp. `Soleil´, Lp. `High Gold´, Lp. `Sunrise´ and Lp. `Vedfire´. To study the propagation of Leucospermum through the simultaneus rooting and grafting, a test was carried out in wich the rootstock Lp. `Spider´ and Lp. pattersonii were compared, these being tolerant to clayey or alkaline-clayey soils, and the comercial cultivars Lp. `Themba´ y Lp. `Raziya´ were used as scions. Additionally, the leaf reduction was studied and the wounding was carried on the rootstock base and the bagging of the graft to favour its maintenance. Both rootstocks were used unrooted to evaluate the concurrent rooted-grafted using the slit graft. The rooting of the stakes was carried out in a well-ventilated umbraclum with 50% reduction in natural light, using a substratum composed by peat-polystyrene 4:6 v/v, with bottom heat (22 ± 2°C) and under a fogging system. A random blocks desing was used with 8 treatments and 3 repetitions, performing 15 grafts per treatment. At the end of the test, Lp. patersonii obtained the best results. Furthermore, it was observed that the rootstock Lp. patersonii with the scion Lp. `Themba´ and whole leaves showed the highest percentage (31%) of transplantable stakes with live scion. With regard to the rooting index, there were no significant differences during the study. El mercado de las proteas se inicia en Canarias en los años 80, desde entonces hasta ahora cuenta ya con unas 60 ha repartidas en las islas de Tenerife, La Palma y Gran Canaria. La producción en las islas se centra sobre todo en el género Leucospermum, siendo los más importantes: Lp. `Succession I´, Lp. `Succession II´, Lp. `Tango´, Lp. `Soleil´, Lp. `High Gold´, Lp. `Sunrise´ y Lp. `Vedfire´. Para estudiar la propagación de Leucospermum mediante el enraizado e injertado simultáneo, se llevó a cabo un estudio en el que se compararon los patrones Lp. `Spider´ y Lp. pattersonii, tolerantes a suelos arcillosos, y como púas se emplearon los cultivares comerciales Lp. `Themba´ y Lp. `Raziya´. Además, se estudió el efecto de la reducción foliar y se llevó a cabo el lesionado en la base del patrón y el embolsado de la púa para favorecer su mantenimiento. Ambos patrones se utilizaron previamente sin enraizar para evaluar el enraizado-injertado simultáneo, utilizándose el injerto de hendidura. El enraizamiento de las estacas se llevó a cabo en un umbráculo ventilado y con una reducción de la luz natural del 50%, empleando un sustrato compuesto por turba y poliestireno 4:6 v/v, con calor de fondo (22 ± 2°C) y bajo un sistema de nebulización. Se empleó un diseño de bloques al azar, con 8 tratamientos y 3 repeticiones, realizándose 15 injertos por tratamiento. Al final del ensayo, Lp. patersonii obtuvo los mejores resultados, además se observa que el patrón Lp. patersonii con la púa Lp. `Themba´ y hojas enteras mostró el mayor porcentaje de estacas trasplantables con púa viva, un 31%. Con respecto al índice de enraizamiento, no hubo diferencias significativas a lo largo del ensayo.

Leucospermum exports from South Africa for the 2013/2014 season comprised 45% (number of stems) of the combined exports of Leucadendron, Leucospermum, and Protea. The determining quality parameter for Leucospermum cut flowers is stem length, with stems sold per centimetre stem length. Over production of short-stemmed Leucospermum cut flowers affect the South African fynbos market share negatively on the export market. Information on the physiology of shoot extension growth, the timing of appendage formation and factors which directly affect shoot length is limited for Leucospermum. Subsequently, shoot extension growth of Leucospermum 'Succession II' (L. lineare × L. cordifolium) and 'Soleil' (L. glabrum × L. cordifolium) was studied to establish its relationship to the timing of appendage formation in two contrasting production areas in South Africa (Hopefield, at 50 m altitude, with particularly warm dry coastal summers; Piketberg, approximately 66 km from Hopefield, but at 800 m altitude, displaying a more moderate, Mediterranean climate). For both regions, shoot extension growth was linear, following an initial lag phase. In all cases 50% or more of the final number of leaves per shoot was differentiated at ca 100 mm stem length. Stems were significantly more extended in the cooler production region of Bo-Piketberg compared to Hopefield and final stem length appears to be area dependent. This data serves as baseline to further studies which aim at evaluating the efficacy of exogenously applied plant growth regulators to increase the stem length of problematic Leucospermum cultivars in warm areas, as well as short-stemmed cultivars such as 'Jelena' produced in cooler areas, in both cases, without compromising flower quality.

Leucospermum cut flower stems are prone to chilling injury and leaf yellowing as a result of long-term cold storage below 4°C as is required when exporting Leucospermum by sea freight from South Africa to European markets. Potted plants of Leucospermum 'Memory' (L. cordifolium selection) and Leucospermum 'Rigoletto' (L. cordifolium × L. glabrum) were subjected to LED lighting (treatments included 'full', 'half' and 'no light' levels) and growth regulator treatments (500 mg L Promalin® and MaxCel™ respectively) to evaluate their efficacy for alleviating cold-induced injuries after a three week cold storage period. The plants subjected to a 'full light' LED treatment showed a significant reduction in chilling injury and leaf yellowing in comparison to plants subjected to the 'no light' treatment. Promalin® and MaxCel™ showed no or little ability to reduce the associated postharvest cold storage disorders. It was concluded that LED at a light energy level of 25.03 ± 2.09 μmol m s was effective at reducing leaf yellowing and chilling injury. A range of Leucospermum cultivars should also be evaluated for its suitability to be placed in cold-storage with supplemented LED lighting.

Seed longevity, the control of dormancy and the eventual fate of seeds were studied in two experiments simulating conditions in natural fynbos. In one experiment a batch of mature, freshly harvested, intact achenes (“seeds”) of the myrmecochorous species Leucospermum cordifolium was divided into lots, of which one was buried in mesic mountain fynbos (experiencing a mediterranean-type climate with hot summers). Other seed lots were stored open at room temperature and in closed nitrogen filled containers at 3 °C, respectively. Stored seeds were sampled and germinated under optimal laboratory (viability estimate) and seed bed (vigour estimate in fynbos) conditions, during autumn, after 0 (control), 1, 2 and 4 years. Low temperature stored seeds maintained a high viability and vigour for c. two years but ambient temperature storage led to a marked decline after 1 year ending in almost complete mortality after 4 years of shelf storage. Four year soil-stored seeds, by contrast, maintained a high viability and vigour, of 80% and 60% respectively of the original seed source values. The soft pericarp/elaiosome in soil-stored seeds disappeared completely whilst the testa became progressively scarified over time. The strongly increased germination rate (velocity) in soil-stored seeds was attributed to natural oxygenation mediated by testa scarification. In another experiment freshly matured intact achenes were oxygenated with 1% H 2 O 2 and the pericarps removed. Disinfection and benzyladenine growth regulator soaking (200 mg L − 1 for 24 h) were applied as separate treatments. The seeds were then sown at 1 cm depth in a standard seed bed in autumn and germination was recorded in the first winter season and, in the undisturbed seed bed, in each subsequent winter for 5 germination seasons. H 2 O 2 oxygenated seeds gave a much higher germination percentage and rate than non-oxygenated seeds, although sporadic germination continued over five germination seasons in all treatments. Seeds germinated only during autumn and early winter each year. The results of the combined experiments suggest that, in nature, Leucospermum seeds can persist underground for long periods. We propose a model of soil-stored seeds in which the intact testa of freshly dispersed seeds is gradually scarified, leading to uneven, extended germination of the young (mainly current season) seed cohort over the first (and possibly several) post-fire germination seasons; and the synchronous germination of the older (scarified), larger, portion of the seed bank, leading to massive species recruitment during the early stages of the first post-fire winter germination season. The numerous wet–dry cycles due to natural rainfall, over prolonged periods in fynbos, may contribute to seed longevity via cellular repair processes.

Ammonia-oxidizing bacteria (AOB) are essential in the biogeochemical cycling of nitrogen as they catalyze the rate-limiting oxidation of ammonia into nitrite. Since their first isolation in the late 19th century, chemolithoautotrophic AOBs have been identified in a wide range of natural (e.g., soils, sediments, estuarine, and freshwaters) and man created or impacted habitats (e.g., wastewater treatment plants and agricultural soils). However, little is known on the plant-species association of AOBs, particularly in the nutrient-starved fynbos terrestrial biome. In this study, we evaluated the diversity of AOBs in the plant canopy of three South African fynbos-specific plant species, namely Leucadendron xanthoconus, Leucospermum truncatulum and Leucadendron microcephalum, through the construction of amoA-gene clone libraries. Our results clearly demonstrate that plant-species specific and monophyletic AOB clades are present in fynbos canopy soils.

Several parameters of flowering stems of two cultivars of protea Leucospermum cordifolium(Knight) Fourc (‘Succession II’ and ‘Tango’) and one of protea Prote L. (Protea susannae x magnifica ‘Susara’) were studied in different commercial plantations. These included length and weight measures, nutrient concentrations of detached parts (flower heads, leaves and stems), and removal of nutrients of harvested flowers. A base for fertilization was also calculated. Harvested flowers of ‘Susara’ removed more P, K and Cu than ‘Succession II’ and ‘Tango’, and more Ca and Fe than ‘Tango’. ‘Succession II’ showed the highest Zn removal. The P and Cu removal by the crop of ‘Susara’ were significantly higher than those of both Leucospermum cultivars, while K, Ca, and Fe outputs of ‘Susara’ exceeded only those of ‘Tango’. The nutrient top removals amounted to 4.55 g m-2 of N, 0.48 g m -2 of P, and 5.26 g m -2 of K. Data as a base to supply fertilizer to each cultivar are given, with N:P:K ratios of 1:0.08:0.87 for ‘Succession II’, 1:0.08:0.83 for ‘Tango’ and 1:0.10:1.27 for ‘Susara’., This study was funded by the Agreement of the High Council of Scientific Research and the Cabildo Insular de La Palma.

The effect of germination incubation regimes, which were chosen on the basis of temperature patterns found within the fynbos (Mediterranean fire-prone shrubland-type) habitat in the Cape Floristic Region during the autumn germinative period, was tested on seeds of 6 Leucospermum species over 5 taxonomic sections ( L. cordifolium , L. cuneiforme , L. erubescens , L. glabrum , L. reflexum , L. vestitum ) . Seeds of each species were scarified, soaked in water and incubated at 15 combinations of diurnal low (16 h) and high (8 h) temperatures. The results indicated that Leucospermum seeds generally require alternating temperatures for germination. Regression analysis of germination percentage responses were used to estimate provisional optimum low [ T O ( LOW )] and optimum high [ T O ( HIGH )] temperature requirements for individual species. These requirements differed markedly among species, and across species they averaged 9.9 and 21.2 °C, respectively. Individual requirements correlated positively with mean late autumn air temperatures obtained from weather stations near the natural habitats of species. We conclude that a diurnal alternating temperature requirement is a character syndrome in Leucospermum in which the T O ( LOW ) and T O ( HIGH ) components are narrowly adaptive, independent and genetically stable characters relating to the known ecophysiological roles of low and high temperature requirements (Brits, Cutting, Brown and Van Staden, 1995). Ecological and adaptive roles of these requirements are compared with those in some persistent small-seeded soil-stored seed banks.

The potted plant industry based on Proteaceae species in South Africa is limited to the domestic markets as protocols for long-term cold storage as required for sea freight to European export markets have not been established. A preliminary investigation was carried out where potted plants of 3 Leucospermum cultivars 'High Gold', 'Raziya' and 'Scarlett Ribbon' were evaluated for chilling injury when stored at 0.5, 4 and 7°C respectively for a 21-day period. 'Scarlett Ribbon' showed the least chilling injury over the entire 10 day evaluation period, irrespective of storage temperature. However, no temperature regime could be recommended as an optimum storage temperature as all plants, irrespective of temperature treatment were considered unmarketable by day 5 of evaluation. In a following trial, potted plants of another 3 Leucospermum cultivars 'Jelena', 'Soleil' and 'Scarlett Ribbon' were evaluated for chilling injury when stored for a 21 day period at 6°C, either in darkness at 100% relative humidity (%RH) as untreated control plants or sprayed with the anti-transpirant Vapor Guard or kept under low light conditions at 60-70% RH. Plants exposed to reduced relative humidity and low light level throughout the storage period displayed no or little chilling injury symptoms and were rated as marketable at the end of the 10 day evaluation period, whereas both the dark-stored control and Vapor Guard- treated plants were considered unmarketable before or on day 5 of evaluation. Significantly lower stomatal conductance was measured within 24 h after removal from storage in potted plants stored in light. Similarly, 'Jelena', the cultivar which had the lowest chilling injury in this trial, also displayed significantly lower stomatal conductance in comparison to 'Scarlett Ribbon' and 'Soleil'. The role of stomata in the incidence of chilling injury during long-term cold storage of potted plants and stem of the members of Proteaceae requires further investigation.

The aim of this study was to assess the growth of Leucospermum ‘Succession II’ (L. cordifolium × L. lineare) and L. ‘Tango’ (L. lineare × L. glabrum) cultivated in La Palma Island (Canary Islands). During the 2008 growing season, stem length measurements were weekly recorded in eight farms, in a range of altitudes from 600 to 867 m, located in most of the production areas of the island. These data were related with environmental conditions to assess if thermal (the growing degree days, the single triangle and the single sine) and photo-thermal models could be used to predict the end of the vegetative growth period of these cultivars. Base temperatures, photo-thermal and thermal units were calculated for both proteas. The model best adapted to ‘Succession II’ was the photothermal one as it showed the lowest coefficient of variation (7.42%), while for ‘Tango’ the lowest coefficient of variation (7.83%) was obtained using the growing degree days model. However, all models gave a good fit to the observed stem length data of both cultivars, although the coefficients of determination of the photo-thermal model were slightly higher than those of the thermal one. In conclusion, both models can be used to predict the date of the end of the vegetative growth of both Proteaceae.

Ants are often considered antagonists when they visit flowers because they typically steal nectar without providing pollination services. Previous research on ant–flower interactions on two species of South African Proteaceae in the Cape Floral Kingdom revealed that the invasive Argentine ant (Linepithema humile), but not native ants, displace other floral arthropod visitors. To determine how common Argentine ant use of inflorescences is, how Argentine and native ant visits differ in the numbers they recruit to inflorescences, and what factors may affect Argentine and native ant foraging in inflorescences, I surveyed 723 inflorescences in 10 species in the genera Protea and Leucospermum across 16 sites and compared ant presence and abundance in inflorescences with abundance at nearby cat food and jam baits. Argentine ants were the most commonly encountered ant of the 22 observed. Argentine ants, as well as six species of native ants were present in all inflorescences for which they were present at nearby baits. Mean Argentine ant abundance per inflorescence was 4.4 ± 0.84 (SE) ants and similar to that of Anoplolepis custodiens and Crematogaster peringueyi, but higher than observed for the other most commonly encountered native ants, Camponotus niveosetosus and Lepisiota capensis. Both Argentine ants and A. custodiens were more likely to be found foraging in spring and under humid conditions, and in inflorescences closer to the ground, with lower sucrose concentrations, and with a greater proportion of open flowers. Argentine ants were more likely to be found in Protea inflorescences, whereas A. custodiens and L. capensis more often visited Leucospermum inflorescences. Considering its displacement of floral arthropods and widespread use of Proteaceae inflorescences, the Argentine ant could be posing a serious threat to plant and pollinator conservation in this biodiversity hotspot.

Grafting is not a common practice in Proteaceae orchards. It does not solve all the growing problems but it can be used to increase the tolerance to some diseases, to get a better adaptation to some soil types, to propagate faster some cultivars/varieties which are difficult to strike from cuttings, to increase production using vigorous rootstocks or to preserve some endangered species. The aim of this study was to evaluate the effect of Leucospermum sp. ‘Spider’ as a rootstock on the vegetative growth and on the yield of four varieties of Leucospermum: ‘Veldfire’, ‘Vlam’, ‘Goldie’ and ‘Memory’. This study was performed in the coastal area of the Alentejo Portuguese region, in 1.5 year old plants that were already thinned. Twenty plants of each variety (ten grafted and ten not grafted) were observed fortnightly. The shoots were counted, their length was registered, and moment of the flower buds emergence was recorded. Grafting seemed to affect positively the rootstock growth on ‘Goldie’. It seemed also to extend the vegetative growing season, especially on ‘Veldfire’. Grafting did not affect either the shoot number or the stem length. The non-grafted plants resisted better to frost.

Development of embryo envelopes in Leucospermum cordifolium (Salisb. ex Knight) Fourcade is presented in a detailed anatomical study, and their structure interpreted ecologically. To support interpretation results for at least six other species of Leucospermum are reviewed, confirming that L. cordifolium is representative of this shrubby group occurring in fynbos, a fire-prone Mediterranean-type vegetation subject to summer drought. The fate of the water-permeable seed coverings subsequent to dispersal is followed in seeds stored experimentally underground. The testa at maturity effectively excludes oxygen in intact soil-stored seeds, thus creating a less common type of physical dormancy which we term ‘anoxia PY’ (oxygen-impermeable physical dormancy). We postulate ‘fire-mediated desiccation-scarification’ testa breakage on a large scale in freshly dispersed (by ants) soil-stored young-seeds, dynamically regulated by drying of different testa layers through fire heat shock (but also by post-fire ambient climate heat), thus alleviating anoxia PY in, and causing undelayed germination of, a major part of the young-seed bank within the ensuing winter germination season. Concurrently a patchy disturbance pattern within a lesser portion of the young-seed cohort causes uneven underground desiccation-scarification by fire, resulting in variable degrees of oxygenation and thus temporally extended (erratic) germination of young-seeds. Both of these strategies are primary long term fitness traits in Leucospermum. By contrast the persistent older-seed cohort, stored underground during the inter-fire period, may become weathered by soil and climate factors (‘soil-mediated abrasion’), which alleviates anoxia PY in old seeds. Thermodormancy of these embryos (physiological dormancy, PD) is enforced in unburnt vegetation by two narrow habitat-dependent daily temperature requirements (~10°C × 20°C), and these have specific ecological functions. The complex Leucospermum system includes balanced adaptive responses to multiple selective pressures. We propose a coherent mechanistic profile for this fynbos genus, synthesising data partly from the literature of other fire-prone ecosystems.

Chilling injury (CI) was recorded in cut flower shoots of Leucospermum 'Gold Dust', 'High Gold', 'Succession II', 'Tango' and 'Vlam' after 24 days storage at 1°C. Cultivars were pulsed with 2% w/v sugar solutions (5 ml/stem) of fructose, glucose, lactulose, mannose or sucrose prior to storage. All sugars significantly reduced CI in 'Gold Dust' and 'High Gold'. Sugar pulsing solutions had no significant effect on reducing CI in either 'Succession II' or 'Vlam' shoots.

Leucospermum 'High Gold' (L. cordifolium x L. patersonii) can be propagated by cutting grafting. To study the effect of wounding of the cuttings (two shallow and opposite incisions), rooting media (coconut fibre and a mixture of peat moss and polystyrene grains) and reduction of the foliar area of the scions on the rooting grafting process, an experiment was carried out in which scions of L. 'High Gold' were wedge grafted onto unrooted cuttings of L. patersonii, which were subsequently rooted with bottom heat (22±2°C) under a fog system. At 6 weeks from planting, the combination of scions with unreduced leaves + unwounded cuttings, rooted in peat moss-polystyrene grains gave 80% of grafted plants. At that time, the mixture of peat moss and polystyrene grains gave a higher quality rooting than the coconut fibre. At the end of the trial, at 16 weeks, all treatments showed grafted plant percentages higher than 70% success. Wounding did not improve rooting. Reduction of the foliar area of the scions did not improve the production of grafted plants.

Leucospermum spp. are indigenous to South Africa and are produced commercially as cut-flowers. The largest proportion of flowers produced in South Africa is exported via air freight to the Netherlands, but also to other European countries. Botrytis cinerea is a damaging post-harvest pathogen on Leucospermum spp. However, with the recent introduction of shipping as a method of transport there has been an increase in post-harvest losses due to spoilage of flowers because the conditions and extended travelling time are more conducive to disease development than air freight. This research project aimed to identify chemical and biological agents for effective control in commercial Proteaceae production. Furthermore, there is a need to identify new chemical and biotic agents for disease management, since many of the pathogens show resistance to the fungicides currently used. Some chemicals are also no longer accepted by the European Union (EU) markets in keeping with EU regulations. During 2006, fungicides with the potential for control of B. cinerea were screened in vitro, and included benomyl, carbendazim + flusilazole, chlorothalonil, cyprodinil + fludioxonil, fenhexamid, iprodione and pyrimethanil. The results of the in vitro test guided selection of fungicides to be evaluated under field conditions. The following fungicides were tested in the field in 2006 at Elsenburg: cyprodinil + fludioxonil, fenhexamid and pyrimethanil. During 2007, two biological control agents (Trichoderma and Bacillus) were also tested under field conditions, as well as alternations between two fungicides (iprodione and fenhexamid) and these biological agents. It was concluded that the fungicides are able to control field infections and that alternations between fungicides and biological agents show potential field control. The biological agents alone were not successful in controlling the pathogen post-harvest.

In 2003, while initiating an investigation into the potential of New Caledonia's native Proteaceae flora for ornamental purposes, we introduced 17 commercial taxa of South African and Australian Proteaceae as cut flowers and foliage in order to diversify the local floriculture industry. These taxa included Leucadendron 'Brook's Red', 'Inca Gold', 'Safari Sunset', 'Wilson's Wonder' and 'Long Tom'; Leucospermum 'Caroline', 'High Gold', 'Purple Gem', 'Scarlet Ribbon' and 'Sunrise'; Protea cynaroides and P. cynaroides 'Australis', Protea laurifolia 'Rose Mink', Protea neriifolia 'Silvertips' and Protea 'Pink Ice'; Serruria florida and Telopea 'Dawn Fire'. An experimental site with supposed adequate pedoclimatic conditions was identified in Northern New Caledonia. Soil analysis showed a silt loam texture with pH 5.6 and a very low level of 7 ppm for plant-available phosphorus. Average temperatures recorded throughout the trial were found to be the coolest (17-18°C) during the months of July and August, with the hottest (25-27°C) months being January and February. Preliminary results indicated that Telopea 'Dawn Fire', Serruria florida, Leucospermum 'Purple Gem' and L. 'Sunrise' were clearly not adapted to the growing conditions of the experimental site. In terms of yield, the Leucospermum cultivars, i.e. 'Scarlet Ribbon', 'Caroline' and 'High Gold' yielded the best results, whilst the two top performing Leucadendron cultivars were 'Inca Gold' and 'Safari Sunset'. Amongst the Protea species, P. neriifolia 'Silvertips' and P. cynaroides showed exceptional production potential.

Proteus, a mythological Greek god, could change into numerous animate or inanimate forms. Carl Linnaeus probably had the remarkable variability within the family in mind when he assigned the name Proteaceae, with more than 330 species located only in the Cape Floral Kingdom. As the domestication of Proteaceae became more sophisticated, the needs in the production and marketing chain increased. A breeding program must also adapt to the needs of the consumer, distributor, flower producer and nurseryman. The ornamental value, suitability for transport and vase life is probably the most important traits to look for in a new ornamental cultivar, whether used as a cut flower or potted plant. The breeding of Fynbos for the past three flowering seasons (2005-2007) focused on the genera Protea and Leucadendron. The breeding process involved hand-controlled and natural open pollination. Hybrids were initially selected on aesthetic value. Most promising breeding lines were then evaluated in different climatic regions of South Africa by participatory producers who suggested new cultivars for release with Plant Breeder's Rights (PBR) protection. The breeding program of the Agricultural Research Council (ARC) has released 20 new cultivars since 1998 through a joint evaluation panel. A further three Protea breeding lines has been submitted for PBR registration in South Africa to address new market trends: 'Sugar Baby' (Protea repens x P. mundii/lacticolor hybrid), 'Sharonet' (Protea eximia × P. susannae hybrid) and 'Lady Pearl' (F2-hybrid derived from a P. magnifica with P. burchelli hybrid).

Although Leucospermum cultivars 'Tango', 'Succession I', 'Scarlet Ribbon', and 'High Gold' have been cultivated in Azores for approximately a decade, no studies have been conducted to assess their growth and developmental patterns. During the 2006-2007 growing season, fortnightly stem length measurements and developmental stages were recorded in farms located on most of Terceira Island's production regions. In this presentation we report the growth and developmental patterns of these four cultivars. We relate these data with environmental conditions and assess the feasibility of thermal and photo-thermal models to predict the average harvest date of these genotypes. We conclude that Leucospermum average harvest date can be predicted with thermal and photo-thermal models, although predictability varies among the cultivars tested in this study.

The objective of this research was to develop in vitro protocols for species of the Proteaceae family. Three model species were selected according to their different growth characteristics: Protea 'Lady Di', Banksia coccinea and Leucospermum 'High Gold'. These three species are considered difficult for vegetative multiplication both ex vitro and in vitro. For Protea 'Lady Di' and Leucospermum 'High Gold' severe pruning was made on stock plants to induce juvenile lateral shoots. The produced buds of Protea were also partially etiolated by different shading systems. Shoots of B. coccinea stock plants were applied with BAP to stimulate the production of lateral shoots in vitro. All the explants were transported in a cooler with their bases in water which demonstrated to be absolutely necessary for the explants to survive in vitro. At the laboratory the explants were defoliated and disinfected using a solution of 5% NaOCl plus nonionic detergent, with different contact times for each species, and two consecutive rinses with distilled sterilised water. The explants were cultivated on MS media supplemented with 30 g L -1 of saccharose and 6 g L -1 of agar, and pH adjusted to 5.7 before sterilisation. The effect of different types and concentrations of plant growth regulators on shoot production, multiplication and induction of adventitious roots were evaluated in L. 'High Gold'. Significant difference was found for the method of partial etiolation in Protea 'Lady Di' depending on the season of the year. In the case of L. 'High Gold' a 90% lateral bud development was recorded when cultivated on a medium supplemented with 1 mg L -1 TDZ. The maximum rate of multiplication for this cultivar was 3.9 compact shoots per explant after four weeks of culture. Shoot elongation of L. 'High Gold' was achieved when explants were subcultured every four weeks on an MS medium supplemented with 1.0 mg L -1 BA for at least four cycles to obtain a rate of multiplication of 2.6. Finally, these buds were cultivated in medium supplemented with different concentrations of IBA (0, 1 and 2 mg L -1 ) to induce adventitious roots, but to date no positive results could be achieved.

The study was carried out during two years in six commercial protea plantations with Leucospermum cultivars 'Scarlet Ribbon', 'High Gold', 'Veldfire', 'Sunrise' and Leucospermum patersonii species located in eight municipalities of La Palma island (Canary Islands). Distribution and behaviour of foliar nutrients Cu, Fe, Mn and Zn across time were studied. Nutritional needs varied in some cases among cultivars and species. Detected levels of Fe and Zn were lower than those previously published. Usually, Cu values and those of Zn were higher at the beginning of the development cycle than at the end of it. The opposite was observed with Fe and Mn. Leucospermum patersonii species showed the highest Fe and Zn concentrations, which suggests a greater requirement of this species for these nutrients. Cultivar 'High Gold' showed the highest concentrations of Cu and 'Scarlet Ribbon' those of Mn. A general stability of micronutrient concentrations was observed in the September, November and/or January samplings, so that November is recommended for taking samples for current foliar analysis. Foliar ranges for the studied nutrients are suggested for these different cultivars and species.

One of the major problems of Proteaceae cultivation in Madeira Island is the high incidence of root diseases caused mainly by Rosellinia necatrix and Armillaria mellea and secondarily by Fusarium solani and Rhizoctonia solani, which lead to the death of plants older than 2 years. The compactness of the soils as well as its richness in organic matter and in other nutrients, together with the climatic conditions of the island (moderate temperatures and a high relative humidity), facilitate the development of root diseases. The aim of this experiment was to study the effect of various treatments in the reduction of mortality of 5-year-old Leucospermum 'High Gold' plants. The treatments consisted of the application to the roots of 1 L of the following solutions: iron sulphate (30 g/L) plus wettable sulphur (10 g/L) (T2 and T6), copper sulphate (25% copper) (20 g/L) with foliar application of Alga-E (15% of algae extracts) (1.5 ml/L) (T3 and T7), commercial bleach (30 ml/L) (T4 and T8), and Talosint (50% cubiet) (10 ml/L) (T5 and T9); the solutions were applied every 3 months (T2 to T5) or every 6 months (T6 to T9). After one year there were no significant differences between treatments, with a plant mortality of 4.8 to 14.3%. After two years, the lowest mortality was obtained with the application of commercial bleach every 6 months (T8) (28.6%); the other treatments gave higher mortality values but with no significant differences between them, except for treatment T5. Treatment T5 was the worst with 66.7% mortality which was statistically different from T4, T8 and T9. None of the treatments were effective in the reduction of the annual plant mortality; treatment T8 gave more consistent results in the two years of trialling period, with 9.5 and 21% for the first and second year, respectively.

The family Proteaceae dominates the nutrient-poor, Mediterranean-climate floristic regions of southwestern Australia (SWA) and the Cape of South Africa. It is well-recognised that mediterranean Proteaceae have comparatively large seeds that are enriched with phosphorus (P), stored mainly as salts of phytic acid in protein globoids. Seed P can contribute up to 48% of the total aboveground P, with the fraction allocated depending on the species fire response. For SWA species, 70-80% of P allocated to fruiting structures is invested in seeds, compared with 30-75% for Cape species, with SWA species storing on average 4.7 times more P per seed at twice the concentration. When soil P is less limiting for growth, seed P reserves may be less important for seedling establishment, and hence plants there tend to produce smaller seeds with less P. For Australian Hakea and Grevillea species the translocation of P from the fruit wall to the seed occurs in the days/ weeks before final fruit dry mass is reached, and accounts for 4-36% of seed P. Seed P content increases with the level of serotiny, though it decreases marginally as a fraction of the total reproductive structure. The greater occurrence of serotiny and higher seed P content within the Proteaceae in SWA supports the notion that SWA soils are more P-impoverished than those of the Cape.

Fusarium wilt of Proteaceae was first reported in 1998 on the genus Protea in the North West province of South Africa and in Zimbabwe. Subsequently it has been reported from the Canary Islands (Tenerife), Chile and Portugal and might also be a potential risk in Australia. Fusarium oxysporum, the causal agent of Fusarium wilt, blocks the vascular tissue of the host plant, thereby causing wilt, chlorosis and plant death. In this study, molecular techniques have been employed to characterise the pathogen, which has only been morphologically identified thus far. Fusarium oxysporum isolates were obtained from Agathosma, Leucadendron, Leucospermum, Protea and Lavandula originating from South Africa, Zimbabwe and Portugal. Partial sequences of the translation elongation factor 1 alpha (EF-1α) gene and the mitochondrial small subunit (mtSSU) region were generated. Phylogenetic analysis of the sequence data confirmed all the isolates to be Fusarium oxysporum. Fusarium oxysporum isolates from a specific host did not cluster together, indicating that isolates from the same host are not clonal. The DNA phylogenies did not follow the formae speciales classification system.

The genus Pilonychiopus Meyer is re-defined and Austrostigmaeus Fan and Beard is relegated to its synonym. Pilonychiopus hallidayi n. sp., the third species in this genus, collected at Cape Town International Airport, South Africa from imported Leucospermum sp. (Proteaceae) originated in Australia, is described. A new genus, Farajimaeus, is created for Pilonychiopus tutus Meyer. A key to the genera and species of the family Stigmaeidae that lack tarsal claws is presented. Measurements of the type specimens of Pilonychiopus cliffortus Meyer and of F. tutus are provided as the original descriptions lacked these data.

A study of nutrient distribution in the flowering stem of proteas was carried out in commercial protea plantations of each of the cultivars ‘Scarlett Ribbon’, ‘High Gold’, ‘Veldfire’, and ‘Sunrise’ of Leucospermum cordifolium, and of the species L. patersonii located in a subtropical zone (La Palma Island, Canarian Archipelago). Flowering stems were cut into different parts: flower bud, leaves from half proximal stem to the flower bud, leaves from half distal stem to the flower bud, half proximal stem to the flower bud, and half distal stem to the flower bud. Nutrient content of the different parts of the flowering stems of the cultivar and the species gave significant differences in some cases, depending on the nutrient and the cultivar. Occasional dissimilarities among the levels of nitrogen (N), phosphorus (P), potassium (K), magnesium (Mg), sodium (Na), copper (Cu), iron (Fe), manganese (Mn), and zinc (Zn) were determined in comparisons among half proximal stems and half distal stems, indepen...

Species of Mycosphaerella and Teratosphaeria represent important foliicolous pathogens of Proteaceae. Presently approximately 40 members of these genera (incl. anamorphs) have been recorded from Proteaceae, though the majority are not known from culture, and have never been subjected to DNA sequence analysis. During the course of this study, epitypes were designated for several important species, namely Batcheloromyces leucadendri, B. proteae, Catenulostroma macowanii, Mycosphaerella marksii, Teratosphaeria bellula, T. jonkershoekensis, T. parva, and T. proteae-arboreae. Several species were also newly described, namely Batcheloromyces sedgefieldii, Catenulostroma wingfieldii, Dissoconium proteae, Teratosphaeria persoonii, T. knoxdavesii, and T. marasasii. Although accepted as being highly host specific, some species were shown to have wider host ranges, such as M. communis (Eucalyptus, Protea), M. konae (Leucospermum, Eucalyptus), M. marksii (Eucalyptus, Leucadendron), T. associata (Eucalyptus, Protea), and T. parva (Eucalyptus, Protea), which in most cases were found to co-occur with other species of Mycosphaerella or Teratosphaeria on Proteaceae. Furthermore, earlier records of T. jonkershoekensis on Proteaceae in Australia were shown to be representative of two recently described species, T. associata and T. maxii. A phenomenon of underdeveloped, or micro-ascospores was also newly observed in asci of T. maculiformis and T. proteae-arboreae. The exact purpose of asci with two distinct types of ascospores remains to be clarified, as both types were observed to germinate on agar.

A study of nutrient removal was carried out in commercial protea plantations of each of the cultivars ‘Scarlett Ribbon,’ ‘High Gold,’ ‘Veldfire,’ and ‘Sunrise’ of Leucospermum cordifolium, and of the species L. patersonii,located in a subtropical region (La Palma, Canarian Archipelago). Flowering stems of optimal commercial quality were taken from each of the mentioned cultivars and species, once a year (2003 and 2004) at harvest time (January to April), and the different organs were analyzed separately. Flowering stems of the cultivars ‘High Gold’ and ‘Veldfire’ presented the highest contents of most studied nutrients, whereas the opposite was observed in ‘Scarlett Ribbon’ and L. patersonii. Mean yields of flowering stems varied very much among cultivars, and the highest ones were those of ‘High Gold’ and ‘Scarlett Ribbon’. The cultivar ‘High Gold’ removed the highest quantities of studied nutrients, and the contrary was observed in L. patersonii for most of them. Suitable rates of nitrogen (N):...

Cheirosporium gen. nov. is characterised by the production of sporodochial conidiomata, semi-macronematous to macronematous onidiophores that possess several distinct sterile branches, and cheiroid, smooth-walled conidia with rhexolytic secession. The 28S rDNA and ITS rDNA operon of this taxon were amplified and sequenced. A BLAST search revealed low homology between Cheirosporium triseriale and existing sequences in public databases, supporting the hypothesis that the species is new to science. Phylogenetic analysis showed that C. triseriale groups with Dictyosporium and allied species, and nests within the Pleosporales (Dothideomycetes, Ascomycota).heirosporium is morphologically distinct from the cheirosporous genera Cheiromyces, Cheiromycina, Dictyosporium, Digitomyces, Digitodesmium and Pseudodictyosporium and these differences are discussed.

The ability of orthodox seeds to survive long-term dry storage is a key prerequisite for ex situ seed conservation in genebanks. However, only a few credible observations of seed survival for ≥ 200 years have been reported. In this paper, seed survival is shown for three species under suboptimal storage conditions for a documented time of ≥ 203 years and carbon dated at 218–270 years. Two species that germinated are in the Fabaceae (Liparia sp. and Acacia sp.). A water-impermeable seed coat, and hence an inability to equilibrate with ambient relative humidity, may contribute to long-term survival of these species. The third species that germinated (Leucospermum sp.) does not have a water-impermeable seed coat, and long-term survival may be linked to an oxygen-impermeable barrier in the seed. These results for species from the Cape Floristic Region, South Africa, suggest adaptation for extreme longevity in seeds of species of seasonally dry, Mediterranean environments.

Proteaceae are adapted to low-nutrient soils in the various regions where they occur. However, harvesting of flowering stems for the cut-flower industry must eventually cause soil nutrient depletion sufficient to reduce yields. Different N forms, and N and P concentrations were supplied to two Proteaceae cultivars ( Leucadendron ‘Safari Sunset’ and Leucospermum ‘Succession’) in a controlled fertigation experiment, and appropriate concentrations for maximum growth with minimum nutrient accumulation or loss were determined. Small additions of N (0.025–0.1 mM) significantly improved growth of both cultivars growing on Strandveld sandy soil. Larger additions of N (up to 2 mM N) resulted in poor growth (both cultivars) and N accumulation in the soil (Safari Sunset). Small additions of P ( ammonium sulphate > calcium nitrate. Phosphorus toxicity symptoms were associated with increased concentrations of leaf P, Ca and Fe. Under conditions of maximum growth (10 μM P and 0.1 mM N) Safari Sunset removed 18 ± 0.6 g N, 1.5 ± 0.1 g P, 5.3 ± 0.6 g K and Succession removed 5.5 ± 0.2 g N, 0.3 ± 0.02 g P, 3.1 ± 0.5 g K over 6 months. At maximum growth, plants acquired more N and P amounts than were supplied, but supplying higher N and P concentrations adversely affected growth. Thus, a more complex or slow-release form of N and P than urea and soluble phosphate, respectively, may provide enough N and P to replace losses from the farm soil at the low concentrations required for proteas.

A study was conducted from November 2000 to February 2001 to establish causes of poor protea plant growth in the Juliasdale and Norton-Darwendale commercial farming areas of Zimbabwe. Soil and leaf samples from farmer-perceived “deficient” and “normal” areas were analyzed. Soils perceived by farmers to be deficient had lower levels of phosphorus (P), potassium (K), calcium (Ca), and magnesium (Mg) compared with the normal soils. In Juliasdale, all cultivars from perceived-deficient samples had lower levels of zinc (Zn) in leaves compared with normal leaves. Leucospermum cultivar ‘Tango’ and Protea cultivar ‘Moonshine’ also had low levels of P and Zn in the deficient plants compared with the normal plants. In Norton-Darwendale, the Protea cultivar ‘Moonshine’ had lower levels of nitrogen (N), P, K, and boron (B) in perceived-deficient samples compared with normal samples. Soil-analysis data were generally comparable to leaf-analysis data, indicating the potential use of leaf analysis to determine ...

7 pages, 4 figures, 5 tables.-- Available online Oct 17, 2005., Foliar samples, SPAD-502 measurements (both along the growing season), and soil samples were taken from protea plants (Leucospermum cordifolium), cultivar ‘High Gold’ cultivated in six commercial plantations of La Palma Island (Canarian Archipelago). Soil data (pH, available P, Ca, Mg, K and Na, and EC) ranged within normal levels for proteas, though O.M. was high. Drainage decreased the potential problem that could be caused by the high clay content of the soils. Leaves showed high concentrations of N, K, Mg, and Na, while Ca and Cu were normal. On the other hand, Fe and Zn presented some low foliar levels, while Mn values fluctuated depending upon the different plantations. N contents were higher in spring, and those of K, Ca, Mg, and Na increased with time. Relative chlorophyll contents (SPAD-502 measurements) were compared with macro- (N, P, Na, K, Ca, Mg) and micro- (Cu, Fe, Mn, Zn) foliar levels. Results showed no positive correlation between leaf chlorophyll and N content. Higher leaf N concentrations were observed in chlorophyll deficient than chlorophyll sufficient leaves at one site and vice versa at another site. In contrast, lower Mn, Fe and Ca concentrations, the last corresponding with higher ratios of N/Ca were observed in chlorophyll deficient than chlorophyll sufficient leaves at several sites. It is concluded that amendment of soils with Mn, Fe and Ca may prevent the development of chlorophyll deficiency symptoms, associated with diminished photosynthetic capacity, in this important commercial cultivar.

The cultivation of proteas for cut-flowers is a thriving industry in Australia and especially Western Australia. However, proteas are susceptible to a ‘sudden death’ syndrome and this has large economic consequences for the industry. Protea plantations in the south-west of Western Australia were assessed during 1999 to 2001 to determine the causes of Sudden Death. Phytophthora cinnamomi was isolated from the stems and roots of dying plants at 11 of the 28 plantations sampled. These included plants from four genera, Leucadendron, Leucospermum, Protea and Serruria. Protea death and decline was also attributed to other fungal pathogens, a range of pests, nutritional disorders and physical factors. The interaction of these factors will be discussed.