Rogério Mamoru Suzuki

Propagation, growth, and carbohydrates of Dendrobium Second Love (Orchidaceae) in vitro as affected by sucrose, light, and dark

In general, plant material grown in vitro has low photosynthetic ability to achieve positive carbon balances. Therefore, a continuous supply of carbohydrates from the culture medium is required, and sucrose has been the most commonly used carbon source. In this paper, we investigate the effects of different sucrose concentrations and the presence and absence of light on the endogenous levels of soluble carbohydrates and starch as well as on the proliferation and growth of Dendrobium Second Love (Orchidaceae) in vitro. The possibility of using etiolated stem segments as a means for micropropagating this hybrid was also verified. The results obtained indicated that the presence and absence of light and the sucrose concentrations used influenced the amounts of soluble carbohydrates and starch and the proliferation of D. Second Love shoots and roots. An increase in sucrose concentration caused a progressive increase in the amounts of total carbohydrates and starch. Under both light conditions, sucrose was the main sugar found in the shoots followed by glucose and fructose. The addition of sucrose to the culture medium up to 2% and 4% was advantageous to the number of shoots produced per explant and the root longitudinal growth in the presence and absence of light, respectively. Shoot and root dry matter and the number of roots formed per explant increased as sucrose concentration was raised up to 6% in both light treatments. The use of dark-grown shoot segments proved to be a useful and reliable alternative for the micropropagation of this hybrid.

Estudo da germinação e crescimento in vitro de Hadrolaelia tenebrosa (Rolfe) Chiron & V.P. Castro (Orchidaceae), uma espécie da flora brasileira ameaçada de extinção

Orchids are seriously endangered species. For that reason, studies regarding their propagation and development are extremely important. Thus, the present study analyzed the influence of the culture medium on the in vitro germination of Hadrolaelia tenebrosa seeds, on the initial development of protocorms until they form seedlings and on seedling growth during the first twelve months in vitro. The results revealed that the ideal methodology for the in vitro culture of Hadrolaelia tenebrosa involves the use the Knudsons C medium for seed germination and seedling initial growth, and the subsequent transfer of six-month-old plants to VW medium where they should remain until acclimatization is carried out. The obtained data showed that the most efficient culture medium for seed germination of that species was not the most adequate for the development of their seedlings. Therefore, the outcomes of such studies are very important for the optimization of propagation techniques for orchid species.

Strong postzygotic isolation prevents introgression between two hybridizing Neotropical orchids, Epidendrum denticulatum and E-fulgens

Abstract Studies on hybrid zones are essential to understand the origin and evolution of reproductive barriers in plants. To achieve this goal, multidisciplinary approaches are often required to investigate the role of multiple reproductive isolation (RI) mechanisms. For Epidendrum denticulatum and E. fulgens, two Neotropical food-deceptive orchid species, we used molecular, cytogenetic and morphological analyses, experimental crosses and environmental envelope models to assess the strength of the RI and the mechanisms that prevent species collapse when hybridization occurs. Based on genetic assignment tests, hybrids between E. denticulatum and E. fulgens were detected. However, the low frequency of hybrid specimens found, coupled with the high morphological differentiation between parental species, suggested that strong barriers exist to interspecific gene exchange. Indeed, hybrid plants were largely sterile, as determined by meiotic data and crossing experiments. In the hybrid zone studied here, strong postzygotic barriers maintain species integrity, and these RI mechanisms may be also important during early stages of speciation.

Germinação e crescimento in vitro de Cattleya bicolor Lindley (Orchidaceae)

A germinacao de sementes de orquideas in vitro vem sendo utilizada desde o inicio do seculo passado. Apesar disso, o conhecimento disponivel a respeito da composicao nutricional dos meios de cultura que favorecem a germinacao e o crescimento in vitro de orquideas ainda e bastante escasso. Diante da ameaca de extincao da Cattleya bicolor e devido a escassez de conhecimento a respeito da germinacao e do crescimento in vitro dessa especie, este trabalho teve como objetivo avaliar a influencia do meio de cultura na germinacao in vitro de suas sementes, no desenvolvimento inicial dos protocormos ate a formacao das plântulas, bem como avaliar o crescimento das plântulas ate os primeiros doze meses de cultivo in vitro. Os resultados mostraram que o meio de cultura ideal para a germinacao in vitro das sementes e para o desenvolvimento inicial das plântulas de Cattleya bicolor e o de Vacin & Went. Recomenda-se que apos seis meses de cultivo as plântulas sejam transferidas para o meio MS ou VW, que promovem o desenvolvimento das plântulas, e mante-las em um destes meios de cultura ate o inicio da aclimatizacao. Tais informacoes sao de grande importância para a otimizacao do processo de propagacao das especies de orquideas nativas do Brasil.

Germinação, crescimento e desenvolvimento in vitro de orquídeas (Cattleya spp., Orchidaceae)

As orquideas no ambiente natural sofrem exploracao devido a sua importância ornamental, levando algumas especies a extincao. O cultivo in vitro e uma forma alternativa para a conservacao ex-situ. Procurou-se determinar um meio de cultura eficiente para a germinacao in vitro de sementes e o crescimento inicial de plântulas de Cattleya forbesii, bem como para o crescimento de plântulas in vitro de Cattleya harrisoniana. No primeiro caso, sementes foram inoculadas em meio de cultura basico de Murashige & Skoog (MS) = T1 e MS basico acrescido de 2,5 g L-1 de carvao ativado (CA) = T2. No segundo, plântulas com 1 ± 0,2 cm de altura foram submetidas aos tratamentos T1, T2, MS com a metade da concentracao original de macro-micronutrientes (T3) e MS com a metade da concentracao original de macro-micronutrientes suplementado com 1,25 g L-1 de CA (T4). Verificou-se aos 30 dias em C. forbesii uma porcentagem de germinacao de 45% em T1 e 90% em T2. A adicao de CA ao meio de cultura trouxe aumento na altura de plântulas de C. forbesii de acordo com analises realizadas aos 180 dias de cultivo. Em relacao ao crescimento de C. harrisoniana, aos 240 dias observou-se que todos os parâmetros medios avaliados (altura da parte aerea, massa de materia fresca total, numero de raizes e folhas, comprimento da maior raiz e diâmetro do pseudocaule) foram significativamente maiores em T2. Dessa forma, sugere-se o uso do meio MS acrescido de 2,5 g L-1 de CA (T2), uma vez que e significativamente favoravel tanto para a germinacao de sementes quanto para o crescimento de ambas as especies.

Effects of light and ethylene on endogenous hormones and development of Catasetum fimbriatum (Orchidaceae)

This study attempted to clarify the effects of dark, light and ethylene on plant growth and endogenous levels of indole-3-acetic acid (IAA), cytokinins and abscisic acid in Catasetum fimbriatum. Dark-incubation fully inhibited root and pseudobulb formation as well as leaf growth, but favored shoot elongation. The results of continuous and active growth in dark-incubated shoots (stolons) were induced by strong apical meristem sink activity and by the significantly increased levels of cytokinins in shoots. In fact, shoot length, cytokinin and IAA levels in dark-incubated shoots were about twice as great as for those grown under light conditions. Moreover, the total cytokinin level in shoots of C. fimbriatum under light conditions without ethylene was significantly higher than that found in roots. High levels of cytokinins in dark-grown stolons may be closely related to the absence of roots in C. fimbriatum. Under light conditions, the increased IAA level in shoots is mediated by ethylene. However, ethylene caused a significant increase of cytokinins in roots of light-treated plants, which may be involved in the retardation of root growth. Since the difference of cytokinins in shoots between ethylene-treated and non-treated plants under light conditions is small, it is concluded that the marked inhibition of leaf growth in ethylene-treated plants can be attributed to ethylene. Zeatin and zeatin riboside are the major cytokinins in C. fimbriatum regardless of the light conditions, ethylene treatment or organ types.

Dark-induced hormone changes coincide with the resumption of light-inhibited shoot growth in Catasetum fimbriatum (Orchidaceae)

Catasetum fimbriatum plants cultivated in the absence of light exhibit continuous shoot growth leading to the formation of nodes and internodes. On the other hand, when these plants are incubated in the presence of light, shoot longitudinal growth is inhibited and pseudobulbs develop just below the shoot apical meristem. These facts provide evidence of a possible influence of light on mitotic cell division in the shoot apex as well as on pseudobulb initiation. The effects of light and dark on the interruption and/or maintenance of shoot apex mitotic activity and the subsequent formation of pseudobulbs in the sub-meristematic regions were investigated by means of histological and hormonal studies. The interruption of shoot apex development occurred around the 150th d of light incubation and seems to have resulted from the establishment of a strong storage sink in the region of the future pseudobulb, in detriment to the continuous activity of the shoot apical meristem. The reduced total cytokinins/IAA ratio in the apex, mainly due to high levels of IAA, could be a key factor in the interruption of cell divisions. Transfer to the dark brings about the resumption of shoot apex development of plants through the re-entrance of cells in the cell cycle which coincides with a significant increase in the total cytokinins/IAA ratio.

Asymbiotic germination, multiplication and development of Alatiglossum fuscopetalum (Orchidaceae) as affected by culture medium, sucrose and growth regulators

Germinacao assimbiotica multiplicacao e desenvolvimento de Alatiglossum fuscopetalum (Orchidaceae) influenciados pelo meio de cultura, sacarose e reguladores de crescimento. Este trabalho teve como objetivo estudar a germinacao e o desenvolvimento in vitro de Alatiglossum fuscopetalum (Hoehne) Baptista. Foram avaliados os efeitos dos meios Murashige/Skoog (MS e ½MS), Knudson (KN), e Vacin/ Went (VW) na germinacao. Analisou-se tambem a infl uencia desses meios na multiplicacao e crescimento de plântulas com 90 dias de idade, e de diferentes concentracoes de sacarose, benziladenina [BA] e acido indolilbutirico [AIB] na multiplicacao e crescimento de plantas com 120 dias de idade. O meio ½MS proporcionou a maior germinabilidade. O meio MS foi superior aos demais em relacao ao crescimento de plântulas de 90 dias, enquanto que o ½MS foi o mais efi caz para a multiplicacao e crescimento de plantas de 120 dias. Concentracoes de 1% a 3% de sacarose favoreceram o crescimento. Para a proliferacao de brotos e crescimento radicial os melhores tratamentos foram 1 mg L-1 de BA e 2 mg L-1 de AIB, respectivamente.