Shoot development in Quercus suber L. somatic embryos

Summary N⁶-Benzyladenine (BA; 0.04–4μM) application to germinated Quercus suber somatic embryos considerably increased caulinar apex elongation frequency and maintained active growth in the plantlets, although it did not have a significant effect on the percentage of shoots with normal morphology. The addition of 0.5 μM indoleacetic acid together with the cytokinin did not have any effect. The use of a low concentration (0.04 μM) of BA allowed the appropriate radicle elongation in all germinating somatic embryos, but higher concentrations arrested this elongation.     

Micropropagation of Rollinia mucosa (JACQ.) baill

Summary A protocol for in vitro propagation of Rollinia mucosa, an important medicinal plant, was developed. The presence of 500 mg l⁻¹ polyvinylpyrrolidone (PVP) during explant excision was important to avoid browning. Axillary buds, adventitious buds, and shoot cluster proliferation were achieved from epicotyl and hypocotyl explants from nursery-grown seedlings. The highest direct organogenesis percentage from hypocotyl explants was obtained upon culture of explants on Murashige and Skoog medium supplemented with 2.2 μM benzyladenine (BA) plus 2.32 μM kinetin. Epicotyl explants display highest regeneration frequency on a medium containing 8.8 μM BA and 0.54 μM naphthaleneacetic acid. Gibberellic acid was necessary for shoot elongation. Root induction was observed when shoots were pretreated with activated charcoal for 7 d in the dark before culture on Woody Plant Medium supplemented with 49.21 μM indolebutyric acid for 10 d. Root development was observed when 20 g l⁻¹ sucrose was used. Rooted plantlets were acclimatized and grown in the greenhouse.     

Effects of ascorbic acid on axillary shoot induction in Tylophora indica (Burm. f.) Merrill.

A procedure for rapid in vitro multiplication of Tylophora indica (Burm. f.) Merrill., an important indigenous medicinal plant, has been developed. Addition of ascorbic acid was essential to induce sprouting of axillary buds. Optimum multiplication was observed on MS medium containing 6-benzylamino purine (5.0 mg l^–1), -naphathalene-acetic acid (0.5 mg l^–1) and ascorbic acid (100 mg l^–1). Rooting of in vitro produced shoots was readily achieved with indole-3-acetic acid alone (1.0 mg l^–1) in MS. The plantlets thus obtained were successfully transferred to pots in large numbers which grew normally.Abbreviations BAP 6-benzylamino purine - 2,4-D 2,4-dichlorophenoxyacetic acid - GA₃ gibberellic acid - IAA indole-3-acetic acid - IBA indole-3-butyric acid - 2ip 2-isopentenyladenine - Kn kinetin - MS Murashige & Skoog media - NAA -naphthalene acetic acid     

Effect of abscisic acid and stratification on somatic embryo maturation and germination of holm oak (<Emphasis Type="Italic">Quercus ilex</Emphasis> L.)

Summary The effect of abscisic acid (ABA) was evaluated during the maturation and germination of holm oak (Quercus ilex L.) somatic embryos. The addition of ABA to the culture medium significantly reduced unwanted recurrent embryogenesis in mature somatic embryos without affecting the germination of embryos subjected to stratification at 4°C. Stratification at 4°C for 2 mo. was the most efficient for stimulating somatic embryo germination of holm oak. The addition of 90 and 450 mM sucrose also improved germination, while higher sucrose concentrations were inhibitory.     

The phylogenetic placement of Ostropales within Lecanoromycetes (Ascomycota) revisited

Results of molecular studies regarding the phylogenetic placement of the order Ostropales and related taxa within Lecanoromycetes were thus far inconclusive. Some analyses placed the order as sister to the rest of Lecanoromycetes, while others inferred a position nested within Lecanoromycetes. We assembled a data set of 101 species including sequences from nuLSU rDNA, mtSSU rDNA, and the nuclear protein-coding RPB1 for each species to examine the cause of incongruencies in previously published phylogenies. MP, minimum evolution, and Bayesian analyses were performed using the combined three-region data set and the single-gene data sets. The position of Ostropales nested in Lecanoromycetes is confirmed in all single-gene and concatenated analyses, and a placement as sister to the rest of Lecanoromycetes is significantly rejected using two independent methods of alternative topology testing. Acarosporales and related taxa (Acarosporaceae group) are basal in Lecanoromycetes. However, if the these basal taxa are excluded from the analyses, Ostropales appear to be sister to the rest of Lecanoromycetes, suggesting different ingroup rooting as the cause for deviating topologies in previously published phylogenies.     

Abscisic acid and ancymidol promote conversion of somatic embryos to plantlets and secondary embryogenesis inAsparagus officinalis L.

Summary The effects of abscisic acid (ABA) (0, 0.09 μM, 0.19 μM, 0.28 μM, and 0.38 μM) or ancymidol (0, 0.98 μM, 1.95 μM, 2.93 μM, 3.90 μM) in embryo germination medium on the conversion of primary embryos to plantlets and secondary embryogenesis were evaluated for asparagus. ABA and ancymidol each significantly enhanced both responses. ABA was more effective than ancymidol in promoting the conversion of primary embryos to plantlets, while the converse was true for the production of secondary embryos. The most effective treatments for embryo conversion were 0.19 and 0.28 μM ABA; 75–77% bipolar and 55–57% globular embryos converted to plantlets. For secondary embryogenesis, the most effective treatments were 1.95 and 2.93 μM ancymidol; 99–101 and 84–86 somatic embryos were produced from 10 globular and 10 bipolar embryos, respectively. Bipolar embryos generally converted to plantlets better than globular embryos, but more secondary embryos were produced from globular embryos than from bipolar embryos in all treatments. ABA and ancymidol also affected the morphology of the plantlets produced. The plantlets from the embryos incubated on the medium with ancymidol had strong and thick shoots and roots, while those on the medium with ABA had long, thin shoots and short thin roots.     

Morphology and development of Nigrosabulum globosum, a cleistothecial coprophile in the Bionectriaceae (Hypocreales)

Recent DNA sequence analyses indicated that Nigrosabulum globosum is a cleistothecial representative of the Bionectriaceae in the Hypocreales, but morphological characters supporting this relationship are unknown. Using light and electron microscopy we followed the development of the ascomata of this species, from the formation of gametangia through to the development of mature ascospores, and observed a series of characters that confirmed its hypocrealean affinities. These included the formation of a gel-filled centrum during early stages of ascoma development, the subsequent appearance of hyaline peridial tissue enclosed within a layer we interpret as representing a melanized uniloculate stroma, apically derived paraphyses, and an ascogenous system that gives rise to asci that were both cylindrical to clavate and globose. Ascospores, previously reported to be smooth, were ornamented with a honeycomb-like reticulum and were able to germinate within the ascoma. The carbonaceous outer (stromatic) walls of the mature, grit-like cleistothecia indicate possible resistance to UV radiation and desiccation. Furthermore, the complement of germinated ascospores would enable mature ascomata to function as propagules that could quickly initiate new growth when transferred to fresh substrate. Our reexamination of N. globosum also provides data that support the hypothesized close relationship with other bionectriaceous, cleistothecial coprophiles, i.e., species of Hapsidospora, and Bulbithecium in particular.     

Foreign exploration for Scirtothrips perseae Nakahara (Thysanoptera: Thripidae) and associated natural enemies on avocado (Persea americana Miller)

Scirtothrips perseae Nakahara was discovered attacking avocados in California, USA, in 1996. Host plant surveys in California indicated that S. perseae has a highly restricted host range with larvae being found only on avocados, while adults were collected from 11 different plant species. As part of a management program for this pest, a “classical” biological control program was initiated and foreign exploration was conducted to delineate the home range of S. perseae, to survey for associated natural enemies and inventory other species of phytophagous thrips on avocados grown in Mexico, Guatemala, Costa Rica, the Dominican Republic, Trinidad, and Brazil. Foreign exploration efforts indicate that S. perseae occurs on avocados grown at high altitudes (>1500 m) from Uruapan in Mexico south to areas around Guatemala City in Guatemala. In Costa Rica, S. perseae is replaced by an undescribed congener as the dominant phytophagous thrips on avocados grown at high altitudes (>1300 m). No species of Scirtothrips were found on avocados in the Dominican Republic, Trinidad, or Brazil. In total, 2136 phytophagous thrips were collected and identified, representing over 47 identified species from at least 19 genera. The significance of these species records is discussed. Of collected material 4% were potential thrips biological control agents. Natural enemies were dominated by six genera of predatory thrips (Aeolothrips, Aleurodothrips, Franklinothrips, Leptothrips, Scolothrips, and Karnyothrips). One genus each of parasitoid (Ceranisus) and predatory mite (Balaustium) were found. Based on the results of our sampling techniques, prospects for the importation of thrips natural enemies for use in a “classical” biological control program in California against S. perseae are not promising.     

Morphology of carrot somatic embryos formed in medium with abscisic acid

Somatic embryogenesis was induced from embryogenie cells derived from cotyledon expiants cultured on MS medium supplemented with 1 mg/L 2,4-D. In order to clarify the effect of abscisic acid (ABA) on the morphology of somatic embryos, embryogénie cell clumps or developing somatic embryos were treated continuously, or briefly, with ABA during culture. When embryogenie cells in MS medium without 2,4-D were treated with 0.04 mg/L ABA for the first week, normal embryos with two cotyledons increased slightly and embryos with anomalous cotyledons decreased. However when cell clumps in 2,4-D-free medium were treated with ABA in the second week normal embryos with two cotyledons decreased prominently and this decrease of normal embryos also occurred in the continuous ABA treatment during culture. Thus the morphological abnormalities in somatic embryogenesis occurred by exogenous ABA treatment beyond globular stage or by continuous treatment. The length of somatic embryos with anomalous cotyledons was larger than that of normal embryos with two cotyledons in control but both the normal and anomalous somatic embryos treated with ABA were almost similar in length. Somatic embryos formed in medium with ABA were larger in size than those in control due mainly to enlarged cotyledons. The enlarged cotyledons were composed of a greater number of cells than those of control. Therefore the enlargement of cotyledon by exogenous ABA seems to be not due to the enlargement of cells in cotyledons.     

Direct and indirect somatic embryogenesis on cotyledon explants of <Emphasis Type="Italic">Quassia amara</Emphasis> L., an antileukaemic drug plant

Summary In vitro propagation of Quassia amara L. (Simaroubaceae) was attempted using mature and juvenile explants. Attempts to establish in vitro culture using leaf and internode explants from a plant more than 15yr old were unsuccessful due to severe phenolic exudation. Plant regeneration through direct and indirect somatic embryogenesis was established from cotyledon explants. Murashige and Skoog (MS) medium with 8.9 μM N⁶-benzyladenine (BA) and 11.7 μM silver nitrate induced the highest number (mean of 32.4 embryos per cotyledon) of somatic embryos. Direct somatic embryogenesis as well as callus formation was observed on medium with BA (8.9–13.3 μM). Semi-mature pale green cotyledons were superior for the induction of somatic embryos. Embryos developed from the adaxial side as well as from the point of excision of the embryonic axis. More embryos were developed on the proximal end compared to mid and distal regions of the cotyledons. Subculture of callus (developed along with the somatic embryos on medium with BA alone) onto medium containing 8.9 μM BA and 11.7 μM silver nitrate produced a mean of 17.1 somatic embryos. Primary somatic embryos cultured on MS medium with 8.9 μM BA and 11.7μM silver nitrate produced a mean of 9.4 secondary somatic embryos. Most of the embryos developed up to early cotyledonary stage. Reduced concentration of BA (2.2 or 4.4 μM) improved maturation and conversion of embryos to plantlets. Ninety percent of the embryos converted to plantlets. The optimized protocol facilitated recovery of 30 plantlets per cotyledon explant within 80d. Plantlets transferred to small cups were subsequently transferred to field conditions with a survival rate of 90%.     

Microorganisms asssociated with Withania somnifera leaves

Microorganisms including bacteria, actinomycetes and fungi were recovered from the leaves of Withania somnifera, which were collected from two altitudinal ranges (0–300 m and 1700–2000 m) in the Asir region, Saudi Arabia. Types and numbers of microorganisms varied according to the altitude and the month of collection. The number of microorganisms was higher on old leaves than that on young ones in most cases. Low altitude exhibited more microorganisms than high altitude. The relationship between meteorological factors and type and number of the recovered microorganisms is discussed. Inoculation of detached healthy leaves of Withania by all recovered fungal species revealed only Alternaria solani as a pathogen of this plant. To confirm pathogenicity, scanning and transmission electron microscopic examination revealed the colonization of this pathogen inside the leaf tissue. Penetration of Withania leaves by the fungus occurred only through stomata, and the invading hyphae were located in the intercellular spaces of leaf tissues. Ultrastructural changes noted in infected cells included changes in chloroplasts and the invagination of the host plasma membrane.     

The effects of mullein plants (Verbascum thapsus) on the population dynamics of Dicyphus hesperus (Heteroptera: Miridae) in tomato greenhouses

The response of Dicyphus hesperus Knight (Heteroptera: Miridae) to whitefly populations in tomato greenhouses was measured in the presence and absence of mullein (Verbascum thapsus L.) as an alternative host plant. The dynamics of the D. hesperus population on tomato (Lycopersicon esculentum Mill.) and on mullein plants were followed through an entire growing season. In houses with mullein plants, more predators occurred on mullein when whitefly density was low on tomato. A mark-release-recapture experiment where rabbit IgG was used as an external marker showed that D. hesperus adults moved from mullein plants to tomato plants. D. hesperus was always more abundant in houses with mullein than in the houses with tomato plants alone. Movements between tomato and mullein plants are discussed as a strategy to optimize predator foraging. The use of mullein as an alternative host plant may contribute to the establishment of D. hesperus and help to preserve the predator population when prey on tomato crops is scarce.     

A new fungal genus, Teracosphaeria, with a phialophora-like anamorph (Sordariomycetes, Ascomycota)

The new genus and species Teracosphaeria petroica is described for a perithecial ascomycete and its anamorph occurring on decayed wood collected in New Zealand. The fungus produces immersed, non-stromatic ceratosphaeria-like perithecia in nature, with hyaline, septate ascospores produced in unitunicate, non-amyloid asci. The anamorph produced in vitro is phialophora-like with lightly pigmented phialides terminating in flaring, deep collarettes that are often noticeably brown with conspicuous periclinal thickening. Phylogenetic analysis of LSU rDNA sequence data indicates that this fungus is distinct from morphologically similar fungi classified in the Chaetosphaeriales, the Trichosphaeriales or the Magnaporthaceae. It forms a monophyletic group with recently described, chaetosphaeria-like ascomycetes, such as the pyrenomycete genus Mirannulata, and shows affinity with the anamorphic species Dictyochaeta cylindrospora. The usefulness of describing anamorph genera for morphologically reduced anamorphs, when anamorph characteristics are actually part of the holomorph diagnosis, is discussed. An apparently contradictory example of the so-called Cordana and Pseudobotrytis anamorphs of Porosphaerella spp. is also discussed.     

Suitability of Aphis fabae, Myzus persicae (Homoptera: Aphididae) and Aleyrodes proletella (Homoptera: Aleyrodidae) as prey for Coccinella undecimpunctata (Coleoptera: Coccinellidae)

The suitability of Aphis fabae Scopoli, Myzus persicae Sulzer and Aleyrodes proletella L. as food sources for Coccinella undecimpunctata L. was evaluated by studying the impact of prey consumption on the predator’s population growth parameters and feeding parameters. Unlike A. proletella, A. fabae and M. persicae supported the development and reproduction of C. undecimpunctata. A. fabae and M. persicae were considered to be essential prey, whereas A. proletella was considered to be an alternative prey. Aphid species showed different degrees of suitability: M. persicae significantly decreased the pre-oviposition period and increased adult longevity, fecundity and fertility compared with A. fabae. Moreover, A. fabae represents a suitable diet for larval development, but is not a suitable food source for adult reproduction. The predator’s population growth parameters, R₀, r_m and λ were increased with M. persicae, whereas T decreased. We found that the 4th instar larvae were the most voracious, particularly when fed on M. persicae; nevertheless, with this prey daily weight gain and feeding efficiency of 4th instar larvae were similar to that of individuals fed with A. fabae.     

Effect of thidiazuron on vegetative tissue-derived somatic embryogenesis and flowering of bamboo Bambusa edulis

Current research on somatic embryogenesis of bamboo uses reproductive tissue as explants. However, it was hard to obtain the explant. Shoots of a local accession (3–4 m high) were used for multiple shoot production. In order to obtain embryogenic callus, nodal and internodal tissues from in vitro plantlets were placed on Murashige and Skoog (MS) medium supplemented with 9.2 M kinetin (KN), 13.6 M 2,4-dichlorophenoxyacetic acid (2,4-D), 0.1% (v/v) coconut milk, and 6% (w/v) sucrose. We studied the effects of sucrose and thidiazuron (TDZ) on callus proliferation. Optimal additives to the MS medium for embryogenic callus proliferation were 0.046 M TDZ, 13.6 M 2,4-D and 3% (w/v) sucrose. TDZ also promoted the germination of bamboo somatic embryos. The germination rate of the somatic embryos exceeded 80% on MS-based medium supplemented with 0.455M TDZ. Naphthaleneacetic acid (NAA) reduced germination. Well-developed plantlets were successfully transferred to soil. There was no albino mutant in subsequent culture. In vitro regenerants and potted plants flowered, but no seeds were produced.     

Abscisic acid and osmotic induction of synchronous somatic embryo development of sweet potato

Summary Somatic embryos of sweet potato have potential as synthetic seeds. The effects of abscisic acid (ABA) (0,0,0.1, 1.0, 10.0 and 50.0 μM) were examined to improve synchrony and proliferation of somatic embryos. Transferring embryos compared to those cultures transferred at day 0. The development of embryos in suspension culture supplemented with ABA was poor. However, when calli proliferation cultures were in gelled medium and pulsed with 0.1 μM ABA for 14 d, the number of somatic embryos increased. Proembryonic masses cultured in mannitol-containing medium (Y=−1.5 MPa) increased embryo development and synchrony of embryo development. Thus, in this work ABA and mannitol have been shown to improve both the total number and the synchrony of sweet potato somatic embryos.