Resumen Los oosporangios y anteridios de Charophyceae son los órganos de reproducción sexual femeninos y masculinos respectivamente. Estas estructuras se caracterizan por su complejidad morfológica y utilidad en taxonomía y sistemática. En el presente trabajo se describen los detalles estructurales y ultraestructurales de la gametogénesis en Chara hydropitys. El material fértil del alga se recolectó en una quebrada tributaria del Río Meléndez en la ciudad de Cali, Colombia (3º21´23´´N - 76º32´5.2´´W). Los especímenes fueron fijados y procesados de acuerdo a los protocolos estándar para la inclusión en resina y obtención de secciones finas que se colorearon con toluidina O (0.3-0.7 μm) para su observación en microscopía fotónica y secciones ultrafinas (60-90 nm) para microscopía electrónica de transmisión (MET). Además, se procesaron muestras para microscopio electrónico de barrido (MEB). Los oosporangios están recubiertos por las células espirales que forman de 10-12 circunvoluciones y terminan en cinco células coronulares. La pared de los oosporangios inmaduros está formada por dos capas que corresponden a la pared de las células espirales y de la oosfera. Al madurar la pared del oosporangio tiene seis capas adicionales, tres de las cuales son aportadas por la oospora y las tres restantes por las células espirales. La oosfera aumenta progresivamente de tamaño a medida que las células espirales crecen y se dividen. En el citoplasma de la oosfera inmadura no se aprecian inclusiones citoplasmáticas conspicuas, pero con la maduración el número de gránulos de almidón aumenta llegando a ocupar la mayor parte del volumen celular. En las células espirales del oosporangio maduro se observan numerosos cloroplastos con prominentes depósitos de almidón entre las lamelas tilacoidales y una vacuola que ocupa casi toda la célula. En las observaciones con MEB se aprecia que la pared externa de la oospora, sobre la zona de la fosa presenta microornamentaciones de tipo verrucado. En los anteridios maduros las células del escudo están fuertemente pigmentadas de color naranja por la presencia de numerosos plastoglóbulos entre las lamelas tilacoidales. De las células del capítulo secundario se desarrollan los filamentos espermatógenos que por divisiones mitóticas unidireccionales y sincrónicas forman los espermatocitos. A partir de estas células haploides por espermiogénesis se desarrollarán los anterozoides biflagelados. Los eventos subcelulares relacionados con estos procesos de división y diferenciación celular incluyen inicialmente cambios en la condensación de la cromatina, pérdida del nucléolo y mayor actividad de los dictiosomas. Posteriormente, el citoplasma se retrae y los orgánulos se alinean a lo largo del núcleo condensado y del aparato flagelar. Los anterozoides maduros emergen a través de un poro lateral de la pared de los espermatocitos. Todos los eventos descritos indican que los procesos de gametogénesis y los detalles estructurales de los gametos son por lo general características ampliamente conservadas en este grupo de algas.

Abstract InCharophyceae, the oosporangia and antheridia are the respective female and male structures of sexual reproduction. These organs are characterized by their morphological complexity and usefulness in taxonomy and systematics. Here we described the structural and ultraestructural details of Chara hydropitys gametogenesis. The fertile material from the algae was collected in a tributary stream of the Río Meléndez in Cali, Colombia (3º21´23´´N - 76º32´5.2´´W) in March 2011. The specimens were fixed and processed following the standard protocols for inclusion in resin. Thin sections (0.3-0.5 μm) were stained with toluidine O, and were observed by photonic microscopy, and additional ultrathin sections (60-90 nm) were observed by transmission electron microscopy (TEM); other samples were processed and observed by scanning electron microscopy (SEM). We found that the oosporangia are covered with spiral cells, forming 10-12 convolutions and ends in five coronula cells. The immature oosporangia wall is formed by two layers that correspond to the wall of the spiral cells and to the oosphere. In mature stages, the oosporangia wall is composed by six additional layers, three of them are provided by the oosphere and the other three are provided by the spiral cells. Oosphere size increases progressively while the spiral cells grow and divide. The cytoplasm of the immature oosphere does not exhibit conspicuous cytoplasmic inclusions, nevertheless, with the maturation, the number of starch granules increases, occupying most of the cell volume. In the spiral cells of the mature oosporangia we observed large number of chloroplast with starch accumulations, between thylakoid lamellae and a vacuole that occupies almost the entire cell. By using SEM it was possible to appreciate, that the external wall of the oospore, more accurately, on the fossa area, shows verrucose micro-ornamentations with verrucae elevations. In mature antheridia, shield cells are strongly pigmented orange due to the presence of a large number of plastoglobules between thylakoid lamellae. The spermatogenous filaments are developed from cells of the secondary capitulum; those, by unidirectional and sincronic mitotic divisions develop the spermatocytes. The biflagellate antherozoids are developed from the haploid cells by spermiogenesis. The subcellular events related with these division and differentiation processes, include first, chromatin condensation, loss of nucleoli and more activity in dictyosomes. Subsequently, retracts the cytoplasm and the organelles are aligned along the condensed nucleus and flagellar apparatus. Mature antherozoids emerge through a side wall pore of the spermatocytes. All the described events showed that the gametogenesis processes and the gametes structural details in general, are widely conserved in this algae group.

Los estudios sobre aspectos reproductivos son escasos en Equisetum. Por eso, hemos realizado un análisis detallado del proceso de esporogénesis, desarrollo de las esporas, ultraestructura de procesos que tienen lugar durante la meiosis, formación de la pared esporal, orbículas y eláteres de E. bogotense, en especímenes procedentes del Cauca, Colombia. Los estudios se efectuaron mediante microscopía fotónica, electrónica de transmisión (TEM) y de barrido (SEM). Los estróbilos llevan numerosos esporangióforos maduros, cada uno con un escutelo peltado, unido al eje del estróbilo por el manubrio y portador de 5-6 esporangios sésiles. Los esporocitos experimentan meiosis dando origen a tétradas de esporas. El tapete pierde la integridad histológica en las primeras etapas de esporogénesis y rodea los esporocitos premeióticos, posteriormente a las tétradas y finalmente las esporas inmaduras, que experimentan cambios citológicos y de tamaño antes de alcanzar la etapa adulta. El esporodermo de las esporas adultas de E. bogotense consiste de seudoendosporio, exosporio y perisporio. Vistos con MEB, el exosporio de las esporas adultas es liso a rugulado con microperforaciones y el perisporio es muriforme, rugado, con pliegues delicados, estrechos, discontinuos, que se distribuyen al azar y delimitan aréolas incompletas. Externamente el perisporio está cubierto por orbículas, que se forman también en la cara externa y los márgenes de los eláteres. Vistos con TEM, el exosporio es una capa de material granular fino y el perisporio, una capa mucho más delgada con orbículas discretas. Los eláteres están formados por dos capas de naturaleza fibrilar, orientadas longitudinalmente y transversalmente. La esporogénesis en E. bogotense es sincrónica, similar a la de E. giganteum, otra especie de distribución tropical que también crece en Colombia.

Studies on some reproductive traits in Equisetum species are scarce and valuable to understand species distribution. Therefore, a detailed study of the sporogenesis process and spore development in E. bogotense is presented, with an analysis of the main events during meiosis, maturation of spores, spore wall ultrastructure, orbicules and elaters. Specimens were collected from 500 to 4 500m in Cauca, Colombia. Strobili at different maturation stages were fixed, dehydrated, embedded in resin, and ultra-microtome obtained sections were stained with Toluidine blue. Observations were made with optical microscopy with differential interference contrast illumination technique (DIC), transmission and scanning electron microscopy (TEM and SEM). Ultrathin sections (70-80μm) for TEM observations were stained with uranyl acetate and lead citrate; while samples for SEM observations, were fixed, dehydrated in 2.2-dimethoxypropane and dried at critical point as in standard methods. Strobili have numerous mature sporangiophores, each one with a peltate structure, the scutellum, bearing five-six sessile sporangia attached to the axis of strobilus by the manubrium. Immature sporocytes (spore mother cells) are tightly packed within the young sporangia. The sporocytes quickly undergo meiosis, by passing the stage of archesporium and give origin to tetrads of spores. The tapetum loses histological integrity during early stages of sporogenesis, intrudes as a plasmodial mass into the cavity of the sporangium, partially surrounding premeiotic sporocytes, and then, tetrads and adult spores. The tapetum disintegrates towards the end of the sporogenesis, leaving spores free within the sporangial cavity. Spores present several cytological changes that allow them to achieve greater size and increase the number of plastids, before reaching the adult stage. Sporoderm includes three layers external to the cytoplasmic membrane of the spore cell, and they are pseudoendospore, exospore and perispore. Viewed with SEM, the exospore is smooth to rugulate, with micro perforations, while the perispore is muriform, rugate, with narrow, delicate, discontinuous, randomly distributed folds delimiting incomplete, irregular areolae, externally covered by of different size, densely distributed orbicules. These orbicules are also found all over the external face and margins of the elaters, while the internal face is smooth and lack orbicules. Viewed with TEM, the exospore is a thick layer of fine granular material, while perispore is a thinner layer of dense, separate orbicules. The elaters are composed by two layers of fibrillar material: an inner layer with longitudinally oriented fibrils and an outer, thicker and less dense layer with fibrils transversely fibrils and abundant, external orbicules. It is suggested that the processes of ontogeny and characters of the sporoderm are relatively constant in Equisetum; however, sporogenesis in E. bogotense is synchronous and this condition has been observed so far only in E. giganteum, a tropical genus also found in Colombia.

Estudios sobre la ontogenia del estróbilo, los esporangios y la biología reproductiva de Equisetum son escasos, por lo tanto, para la especie E. giganteum, se estudiaron estos aspectos en especímenes recolectados a orillas del Río Frío, Santander, Colombia (2 200m). Los estróbilos en diferentes etapas de maduración fueron fijados, deshidratados, embebidos en parafina, seccionados en micrótomo rotatorio y teñidos con safranina O-fast green. Las observaciones se efectuaron mediante un microscopio óptico de alta resolución con contraste diferencial de interferencia (DIC) y microscopio de fluorescencia. Los estróbilos se inician a partir del meristemo apical, tanto en el eje principal como en los laterales, sin diferencias en el proceso de ontogenia y esporogénesis entre estróbilos de diferentes ejes. Sucesivas mitosis y diferenciación celular conducen al crecimiento del estróbilo, y a la formación de los esporangióforos peltados, formados por el manubrio, o porción basal con aspecto de pedicelo, el escutelo, o porción apical aplanada y las iniciales del esporangio, los cuales se diferenciarán para formar la pared del esporangio, los esporocitos y el tapete. No se forma arquesporio y los esporocitos experimentan meiosis para formar tétradas de esporas. El tapete mantiene la integridad histológica hasta la formación de las tétradas y en esa etapa forma un plasmodio que invade la cavidad esporangial la cual rodea parcialmente las tétradas y luego las esporas, y aparecen las cámaras plasmodiales, un término propuesto aquí para las formaciones designadas en inglés "tapetal gaps". La pared del esporangio queda reducida a dos capas celulares: una externa con engrosamientos lignificados en todas las paredes celulares y una interna picnótica. Al finalizar la esporogénesis, el tapete degenera, y las esporas, con exosporio, perisporio delgado, casi membranáceo y eláteres quedan libres en la cavidad esporangial. El esporodermo, los núcleos y nucléolos presentan fluorescencia roja, inducida por coloración con safranina O, mientras que los eláteres y las células de la pared del esporangio presentan autofluorescencia amarillo-naranja.

Ontogeny of strobili, sporangia development and sporogenesis in Equisetum giganteum (Equisetaceae) from the Colombian Andes. Studies on the ontogeny of the strobilus, sporangium and reproductive biology of this group of ferns are scarce. Here we describe the ontogeny of the strobilus and sporangia, and the process of sporogenesis using specimens of E. giganteum from Colombia collected along the Rio Frio, Distrito de Sevilla, Piedecuesta, Santander, at 2 200m altitude. The strobili in different stages of development were fixed, dehydrated, embedded in paraffin, sectioned using a rotatory microtome and stained with the safranin O and fast green technique. Observations were made using differential interference contrast microscopy (DIC) or Nomarski microscopy, an optical microscopy illumination technique that enhances the contrast in unstained, transparent. Strobili arise and begin to develop in the apical meristems of the main axis and lateral branches, with no significant differences in the ontogeny of strobili of one or other axis. Successive processes of cell division and differentiation lead to the growth of the strobilus and the formation of sporangiophores. These are formed by the scutellum, the manubrium or pedicel-like, basal part of the sporangiophore, and initial cells of sporangium, which differentiate to form the sporangium wall, the sporocytes and the tapetum. There is not formation of a characteristic arquesporium, as sporocytes quickly undergo meiosis originating tetrads of spores. The tapetum retains its histological integrity, but subsequently the cell walls break down and form a plasmodium that invades the sporangial cavity, partially surrounding the tetrads, and then the spores. Towards the end of the sporogenesis the tapetum disintegrates leaving spores with elaters free within the sporangial cavity. Two layers finally form the sporangium wall: the sporangium wall itself, with thickened, lignified cell walls and an underlying pyknotic layer. The mature spores are chlorofilous, morphologically similar and have exospore, a thin perispore and two elaters. This study of the ontogeny of the spore-producing structures and spores is the first contribution of this type for a tropical species of the genus. Fluorescence microscopy indicates that elaters and the wall of the sporangium are autofluorescent, while other structures induced fluorescence emitted by the fluorescent dye safranin O. The results were also discussed in relation to what is known so far for other species of Equisetum, suggesting that ontogenetic processes and structure of characters sporoderm are relatively constant in Equisetum, which implies important diagnostic value in the taxonomy of the group. Rev. Biol. Trop. 59 (4): 1845-1858. Epub 2011 December 01.

A new trypanosomatid species, Blastocrithidia cyrtomeni, is herein described using morphological and molecular data. It was found parasitising the alimentary tract of the insect host Cyrtomenus bergi, a polyphagous pest. The morphology of B. cyrtomeni was investigated using light and transmission microscopy and molecular phylogeny was inferred from the sequences of spliced leader RNA (SL rRNA) - 5S rRNA gene repeats and the 18S small subunit (SSU) rRNA gene. Epimastigotes of variable size with straphanger cysts adhering to the middle of the flagellum were observed in the intestinal tract, hemolymph and Malpighian tubules. Kinetoplasts were always observed anterior to the nucleus. The ultrastructure of longitudinal sections of epimastigotes showed the flagellum arising laterally from a relatively shallow flagellar pocket near the kinetoplast. SL RNA and 5S rRNA gene repeats were positive in all cases, producing a 0.8-kb band. The amplicons were 797-803 bp long with > 98.5% identity, indicating that they originated from the same organism. According to the sequence analysis of the SL-5S rRNA gene repeats and the 18S SSU rRNA gene, B. cyrtomeni is different from all other known species or isolates of Trypanosomatidae. Both analyses indicate that among known species, it is most closely related to Blastocrithidia triatomae.

Se evaluaron las características morfológicas de siete variedades de harina nativa de yuca y polvillo de fique. Las muestras fueron dispersadas en albúmina de huevo, posteriormente extendidas sobre placas de vidrio y teñidas con azul de toluidina para su observación al microscopio. Se usó la técnica de Microscopía Óptica de Alta Resolución (MOAR) por contraste diferencial de interferencia (DIC), para observar las muestras y caracterizar las imágenes. Se encontraron en las harinas gránulos de almidón esféricos y semiesféricos algunos con formas truncadas, y estructuras fibrosas de diferentes formas. En el polvillo de fique se observaron microfibrillas longitudinales en forma de cintas, celdas y espirales. Este estudio permitió caracterizar la morfología de las materias primas estudiadas, cuya información es punto de partida para la continuidad de su uso en el campo de los materiales biodegradables.

Morphological characteristics of seven varieties of native cassava flour and fique dust were evaluated. The samples were dispersed in egg albumen, then spread on glass plates and stained with toluidine blue for observation under the microscope. High Resolution Light Microscopy technique (HRLM) was used, with differential interference contrast (DIC) to observe the samples and characterize the image. Spherical and hemispherical starch granules, truncated and some with fibrous structures in different ways, were found in flour. In fique dust longitudinal microfibrils were observed in the form of tapes, cells and spirals. This study allowed to characteríze the morphology of the raw materials studied, whose information is the starting point for its continued use in the field of biodegradable materials.

Nós avaliamos as características morfológicas de sete variedades de farinha de mandioca nativas e pó de fique. As amostras foram dispersas em albúmina de ovo, em seguida, espalhar sobre placas de vidro e corados com azul de toluidina para observação ao microscópio. Utilizamos a técnica de Microscopia de Alta Resolução de Luz (HRLM) com contraste de interferência diferencial (DIC) de observar as amostras e caracterizar as imagens. Foram encontradas no amido de farinha de grãos esféricos e hemisférica, truncada e com algumas estruturas fibrosas de diferentes maneiras. Em microfibrilas poeira fique longitudinal foram observados sob a forma de fitas, e as células em espiral. Este estudo permitiu caracterizar a morfología das materias-primas estudadas, cuja informação é o ponto de partida para a sua utilização continuada na área de materiais biodegradáveis.