Se diseñó y caracterizó un sistema reportero para ser capturado por anticuerpos enlazados a placas de ELISA. El sistema fue diseñado con una proteína altamente antigénica y específica, la rK346 de Leishmania infantum. La rK346 fue acoplada a la peroxidasa C de rábano picante (HRPc) de Armoracia rusticana usando glutaraldehido o sulfo-SMCC. La conjugación con glutaraldehido fue realizada en dos pasos. La separación de los conjugados fue llevada a cabo a través de una cromatografía de exclusión molecular sefarosa S-200 (CES), las fracciones fueron analizadas midiendo la actividad HRPc y por placas ELISA sensibilizadas con inmunoglobulina G policlonal anti-rK346, purificada desde suero de conejo. Se obtuvo una población heterogénea de conjugados rK346-HRPc en un rango de pesos moleculares entre 109,7 ± 16,5 a 67,6 ± 10,1 kDa; con estequiometria rK346-HRPc de 1:2; 2:1; 3:1; y 2:2. La conjugación usando sulfo-SMCC se llevó a cabo primero introduciendo grupos -SH en la HRPc usando el reactivo SATA; el antígeno se modificó con sulfo-SMCC. La separación y el análisis de los conjugados se realizaron de forma similar que con el glutaraldehido, resultando en una población heterogénea de conjugados rK346-HRPc con un rango de pesos moleculares entre 150,5 ± 22,6 a 80,0 ± 12,0 kDa; con estequiometria rK346-HRPC de 2:1; 1:2; 2:2 y 1:3, y con una eficiencia de conjugación incrementada en comparación con glutaraldehido. De esta forma, se habilitó al sulfo-SMCC como un reactivo potencial para acoplar antígenos a la HRPc, como método para el diseño de un sistema reportero económico, especifico y fácil de aplicar, útil en la evaluación de individuos en riesgo y/o en estados tempranos o avanzados de leishmaniasis visceral.

It was designed and characterized a reporter system to be captured by antibodies bound to ELISA plates. The system was designed with the rK346 from Leishmania infantum, a highly antigenic and specific protein. The rK346 was coupled to the horseradish peroxidase C (HRPc) from Armoracia rusticana using glutaraldehyde or sulfo-SMCC. Glutaraldehyde conjugation was performed in two steps. Separation of conjugates was carried out using a Sepharose S-200 in size exclusion chromatography (SEC); fractions were analyzed via HRPc activity and through ELISA plates sensitized with polyclonal anti-rK346 IgG purified from rabbit serum. A heterogeneous population of conjugates rK346-HRPc was obtained with molecular weights ranging between 109.7 ± 16.5 to 67.6 ± 10.1 kDa; with rK346-HRPc stoichiometries of 1:2; 2:1; 3:1; and 2:2. Conjugation using sulfo-SMCC was carried out first by introducing -SH groups onto the HRPc using the SATA reagent and the antigen was modified with sulfo-SMCC during 45 min. Separation and analysis of conjugates was performed similarly as with glutaraldehyde, resulting in a heterogeneous population of conjugates rK346-HRPc with molecular weights between 150.5 ± 22.6 to 80.0 ± 12.0 kDa; with rK346-HRPC stoichiometries of 2:1; 1:2; 2:2; and 1:3, with an increased conjugation efficiency in comparison with glutaraldehyde. This enables sulfo-SMCC to be used as a potential reagent for coupling the antigen to the HRPc, to design an economic, specific and easy method to apply as a reporter system, available to assess individuals at risk and/or at early and late stages of visceral leishmaniasis.

Eucalypt plantations in Venezuela occupy 14% of land covered by savannas. Since water use by plantations in the savannas of Mapire (Venezuelan Llanos) was apparently similar to or lower than in the natural vegetation, evaluating how carbon fixation by plantations compares to the natural vegetation could help to elucidate whether plantations have higher water-use efficiency (WUE) than the savanna. We aimed to determine, using measurements of leaf gas exchange, the seasonal changes in photosynthetic rate (P N) and WUE of a seasonally dry savanna in Mapire, composed mainly of the grass Trachypogon vestitus and the tree Curatella americana, and trees of Eucalyptus urophylla growing in an experimental plantation. Daily integrated WUE (WUEint) was highest in C. americana and similar between T. vestitus and E. urophylla during both seasons. The Gross carbon fixation (GCF) at the leaf level in T. vestitus was similar to C. americana and lower than E. urophylla during the dry season, whereas during the rainy season it was nearly as high as in E. urophylla and both higher than C. americana. Gross carbon fixation (GCF) in T. vestitus was higher than in C. americana, but lower than in E. urophylla during the dry season, while it was much higher than the other two species during the rainy season. The contribution to savanna GCF of C. americana was small and constant throughout the seasons. The GCF in eucalypts was 15% higher than in the savanna during the dry season, but 45% lower during the rainy season, GCF averaging 85% of that in the savanna. Since water use by plantations is lower than in the savanna, they have a large potential for C sequestration without significantly altering ground water availability.

Nicotiana glauca, a shrub native to southern South America, is widely distributed in the Americas, from Patagonia to the USA, from zero up to 3,700 m; it rapidly invades disturbed environments. In Venezuela, it has been reported from zero up to 2,000 m growing in contrasting conditions of relative humidity, temperature, rainfall, and salinity. In order to gain insight into the extent and mode of resistance to drought and salinity in N. glauca, we studied the effect of these factors on water relations and photosynthesis under both natural and greenhouse conditions. In the field, water potential, photosynthetic rate (A) and stomatal conductance (gs), but not relative water content (RWC) decreased because of drought. Manual removal of epicuticular wax increased excess energy dissipation through non-photochemical quenching without altering the capacity of photochemical quenching. In the greenhouse, water deficit as well as salinity resulted in osmotic adjustment; at the end of the experiment, turgor potential and RWC under water deficit were similar to control and higher under salinity. Water deficit and salinity caused marked decreases in A and gs. There were very few or no changes with natural drought, salinity or experimental water deficit in potential quantum efficiency of PSII, which could be explained partly by an increased non-photochemical quenching. We conclude that the partial tolerance to drought and salinity in plants of N. glauca resides in their ability to achieve water conservation through stomatal closure and osmotic adjustment, reduce absorption of excess radiation through the presence of leaf wax and dissipate it through increased non-photochemical quenching. All these characteristics confer plants of N. glauca advantages to invade disturbed areas, subject to salinity and/or seasonal drought under high irradiance.

Las especies del género Lantana son conocidas por el alto valor comercial de sus aceites naturales, pero se sabe poco de su respuesta fotosintética a variables ambientales. El efecto del déficit hídrico sobre la actividad fotosintética se estudió en tres especies de Lantana: L. caracasana, L. canescens y L. camara, con tratamientos de suspensión del riego. L. caracasana tuvo la mayor asimilación de carbono durante el período de medida y el déficit en la disponibilidad de agua ocasionó una disminución del potencial hídrico, de la fotosíntesis y de la conductancia estomática en las tres especies. Los cambios observados en las variables de fluorescencia, en la eficiencia cuántica aparente y en la fotosíntesis a luz saturante, sugieren una regulación descendente de la actividad fotoquímica del fotosistema II durante el déficit hídrico. En plantas control se observaron pocas diferencias entre especies en la respuesta de la fotosíntesis a la concentración intercelular de CO2. La temperatura óptima para la fotosíntesis varió entre 25-26ºC. El efecto del déficit hídrico sobre el intercambio gaseoso de las tres especies fue diferente, siendo L. camara la más afectada.

The species of the Lantana genus are known by the high commercial value of its natural oils, but their photosynthetic response to environmental variables is not well known. To characterize the photosynthetic capacity and establish the effect of water deficit on photosynthesis, plants of three Lantana species (L. caracasana, L. canescens and L. camara) were submitted to water deficit by holding watering.The highest diurnal carbon assimilation was observed in L. caracasana and water deficit caused a decline in water potencial, photosynthesis and stomatal conductance of all species. Changes observed in variable fluorescence, apparent quantum yield and photosynthesis at saturated light, suggested a down regulation of photochemical activity of photosystem II during water deficit. In well irrigated plants, slight differences were observed among species in the photosynthetic response to intercellular CO2 concentration. Optimum temperature of photosynthesis was within 25-26ºC. The effect of water deficit on the photosynthetic parameters varied with the species, being L. camara the most affected.

Nas árvores da floresta sazonalmente inundada do rio Mapire em Venezuela, a inundação cedo induz uma diminuição reversível na condutância foliar e na taxa fotossintética. Com o objetivo de encontrar uma explanação anatômica para as respostas observadas na trocas gasosas nas folhas, as características das folhas emersas desenvolvidas sob a drenagem ou após três meses de inundação foram examinadas nas espécies nas espécies arbóreas Acosmium nitens, Campsiandra laurifolia, Duroia fusifera, Eschweilera tenuifolia Pouteria orinocoensis e Symmeria paniculata, e nas folhas desenvolvidas somente sob inundação de Inga spuria e Tachigali davidsei. A anatomia foi notavelmente similar entre as espécies e as famílias e consistiu em um parênquima paliçádico bi-celular, um parênquima lacunoso de 5-6 células e grande espessura da folha inteira. A anatomia assemelhou-se também àquele das xerófitas ou das espécies sempre-verdes possuindo cutículas grossas, células epidérmicas grandes, paredes anticlinais das epidermes engrossadas e uma abundância de esclerênquima. Folhas das árvores inundadas v. não inundadas não eram qualitativamente diferentes. A área foliar específica assemelhou-se a valores de espécies decíduas em florestas secas tropicais. Nenhuma diferença quantitativa foi encontrada entre as folhas desenvolvidas em árvores não inundadas e inundadas, à exceção de uma redução na espessura da folha inteira do E. tenuifolia, P. orinocoensis e S. paniculata e uma mudança na contribuição do parênquima paliçádico a espessura foliar em E. tenuifolia. O tamanho e a densidade estomáticos nestas espécies exclusivamente hipostomáticas permanesceram não afetados pela inundação. Uma diminuição sob inundação na espessura foliar pode estar relacionado ao aumento da condutância mesofilar e conseqüentemente da taxa fotossintética.

In trees of the seasonally flooded forest of the Mapire River in Venezuela, early flooding induces a reversible diminution in leaf conductance and photosynthetic rate. With the aim of finding an anatomical explanation for the observed responses of leaf gas exchange, the characteristics of emerged leaves developed under drainage or after three months of flooding were examined in the tree species Acosmium nitens, Campsiandra laurifolia, Duroia fusifera, Eschweilera tenuifolia, Pouteria orinocoensis and Symmeria paniculata and in leaves developed only under flooding in Inga spuria and Tachigali davidsei. Anatomy was remarkably similar among species and families and consisted of a bi-layered palisade parenchyma, a 5-6-cell-thick spongy parenchyma and large whole-leaf thickness. Anatomy also resembled that of xerophytes or evergreen species by possessing thick cuticles, large epidermal cells, thickened anticlinal epidermal cell walls and an abundance of sclerenchyma. Leaves of flooded v. un-flooded trees were not qualitatively different. Specific leaf area resembled values of deciduous species in tropical dry forests. No quantitative differences were found between leaves developed in un-flooded and flooded trees, with the exception of a reduction in whole-leaf thickness of E. tenuifolia, P. orinocoensis and S. paniculata and a change in the contribution of palisade parenchyma to leaf thickness in E. tenuifolia. Both stomatal size and density in these exclusively hypostomatous species remained unaffected by flooding. A decrease under flooding in whole-leaf thickness may have resulted in an increase in mesophyll conductance and therefore photosynthetic rate.

Se realizaron medidas del estado hídrico, intercambio gaseoso y características anatómicas foliares en dos especies (Brownea coccinea y Spatiphylum cannifolium) que crecen naturalmente a lo largo de un gradiente natural de CO2, desde una concentración supra-atmosférica (CS) de 35000µmol·mol-1 a una concentración ambiental (CA) de 435µmol·mol-1, en el bosque ribereño del río Santa Ana (Estado Sucre, Venezuela). Las medidas se realizaron en dos épocas contrastantes, lluvia y sequía. El potencial hídrico (psi) fue cerca de 60% mayor en lluvia que en sequía en ambas especies. El crecimiento en CS no ocasionó cambios en el psi de B. coccinea, mientras que en S. cannifolium causó un descenso, estos cambios estuvieron acompañados en ambas especies con disminuciones en el potencial osmótico (<FONT FACE=Symbol>y p</FONT> ) Las elevadas concentraciones de CO2 causaron una mejora en la fotosíntesis (A) y la conductancia estomática (g s) de B. coccinea, pero en S. cannifolium no se encontraron diferencias significativas. En las plantas de ambas especies que crecen en CS las hojas eran 10% más gruesas debido a un incremento del grosor de los tejidos del mesófilo en la misma proporción; además, las plantas de S. cannifolium que crecen en CS tiene lugar una mayor deposición de ceras epicuticulares durante la época de sequía.

In order to gain knowledge on the physiological and anatomical responses of plants to long-term growth under elevated CO2 concentrations, measurements of water status, leaf gas exchange and leaf anatomical characteristics were made during the rainy and the dry season in plants of Brownea coccinea and Spatiphylum cannifolium growing along a natural CO2 gradient, from 35000µmol·mol-1 (supra-atmospheric CO2 concentration, SC) to 435µmol·mol-1 (ambient CO2, AC) in the riparian forest of the Santa Ana river, Sucre State, Venezuela. Water potential (psi) was higher during the rainy than the dry season in both species. Growth under SC did not cause changes in psi of B. coccinea, whereas it decreased it in S. cannifolium; both drought and growth under SC decreased leaf sap osmotic potential (<FONT FACE=Symbol>yp</FONT>) in both species, helping maintain turgor pressure in B. coccinea but not in S. cannifolium. Elevated CO2 increased photosynthetic rate (A) and leaf conductance (g s) of B. coccinea under drought but no effect was found in S. cannifolium. Leaves of both species growing under SC were 10% thicker than under AC, owing to a proportional increase in the thickness of the mesophyll tissues; additionally, during the dry season a higher deposition of epicuticular waxes took place on leaves of S. cannifolium growing under SC. Changes in photosynthetic rate, leaf conductance and leaf anatomy suggest acclimation to growth under elevated CO2.

Realizaram-se medidas do estado hídrico, intercambio gasoso e características anatômicas foliares em duas espécies (Brownea coccinea e Spatiphylum cannifolium) que crescem naturalmente ao longo de um gradiente natural de CO2, desde uma concentração supra-atmosférica (CS) de 35.000mmol·mol-1 a uma concentração ambiental (CA) de 435mmol·mol-1, no bosque marginal do rio Santa Ana (Estado Sucre, Venezuela). As medições se realizaram em duas épocas contrastantes, chuva e seca. O potencial hídrico (Y) foi aproximadamente 60% maior na chuva que na seca em ambas espécies. O crescimento em CS não ocasionou mudanças no Y de B. coccinea, enquanto que no S. cannifolium causou um descenso, estas mudanças estiveram acompanhadas em ambas espécies com diminuições no potencial osmótico (Yð?) As elevadas concentrações de CO2 causaram uma melhora na fotossínteses (A) e a condutância estomática (g s) de B. cocinea, mas em S. cannifolium não se encontraram diferenças significativas. Nas plantas de ambas espécies que crescem em CS as folhas eram 10% mais grossas devido a um incremento na espessura dos tecidos do mesófilo na mesma proporção; além disso, nas plantas de S. cannifolium que crescem em CS acontece uma maior deposição de ceras epicuticulares durante a época da seca.

La gran cantidad de información acerca de los efectos de un incremento experimental de la [CO2] de cultivo sobre la fisiología de plantas de zonas templadas contrasta con la reducida información disponible para plantas tropicales. En atención a ello, se estudiaron las respuestas fotosintéticas a [CO2] elevadas de plantas que crecen en Venezuela. Se cultivaron plantas de cuatro especies xerófitas bajo una [CO2] elevada, para elucidar el efecto de ésta sobre la fotosíntesis y el uso de agua de especies que normalmente enfrentan déficits hídricos. La fotosíntesis aumentó alrededor de 3,5 veces y la [CO2] elevada retrasó la disminución de la tasa fotosintética en sequía. Dado que en muchos estudios se ha encontrado que el estímulo inicial de la fotosíntesis por una [CO2] elevada desaparece en el tiempo debido a las limitaciones de sumidero de asimilados, hicimos un estudio del efecto de la [CO2] elevada sobre la fotosíntesis de plantas de yuca cultivadas sin limitaciones de suelo, encontrando que la tasa fotosintética no disminuyó durante todo el ciclo de cultivo. La existencia de un sumidero grande (la raíz) unida a una gran disponibilidad de sustrato, permitió que la estimulación de la tasa fotosintética por la [CO2] elevada continuara en el tiempo. Se evaluó el efecto de [CO2] muy altas sobre la fotosíntesis de plantas silvestres que crecen alrededor de emanaciones naturales, encontrándose que tales concentraciones no sólo no inhiben la tasa fotosintética, sino que la promueven muy por encima de la de los controles.

The large amount of information on the effects of an experimental increase in [CO2] on the physiology of plants in temperate zones is in contrast with the scant information available on tropical plants. In view of this, the photosynthetic responses to elevated [CO2] of plants growing in Venezuela were studied. In order to elucidate the effect of elevated [CO2] on the photosynthesis and water use of species that are normally subjected to water deficit, plants of four xerophytic species were grown under elevated [CO2]. Photosynthesis increased approximately 3.5 times and elevated [CO2] delayed the decrease of photosynthetic rate under drought. Since in many studies the initial promotion of photosynthetic rate caused by elevated [CO2] has been observed to disappear in time due to limitations imposed by the sinks, we studied the effect of elevated [CO2] on the photosynthesis of cassava plants grown without soil limitations. Photosynthetic rate did not decrease at any time during the growth cycle of the crop. The occurrence of a large sink (the root) together with a sufficiently large volume of substrate permitted the maintenance of a higher photosynthetic rate due to elevated [CO2]. The effect of very high [CO2] on photosynthesis was evaluated in wild plants growing near natural sources; such high [CO2] did not inhibit the photosynthetic rate and promoted it well above that of controls.

A grande quantidade de informação sobre os efeitos de um incremento experimental do [CO2] de cultivo sobre a fisiologia de plantas de zonas temperadas contrasta com a informação reduzida disponível para plantas tropicais. Em atenção a isso, estudaram-se as respostas fotosintéticas ao [CO2] elevados de plantas que crescem na Venezuela. Foram cultivadas plantas de quatro espécies xerófitas sob um [CO2] elevado, para elucidar o efeito desta sobre a fotosíntese e o uso de água de espécies que normalmente enfrentam déficits hídricos. A fotosíntese aumentou ao redor de 3,5 vezes e o [CO2] elevado atrasou a diminuição da taxa fotosintética em seca. Dado que em muitos estudos foi encontrado que o estímulo inicial da fotosíntese por um [CO2] elevado desaparece no tempo devido às limitações de sumideiro de assimilados, fizemos um estudo do efeito do [CO2] elevado sobre a fotosíntese de plantas de mandioca cultivadas sem limitações de solo, encontrando que a taxa fotosintética não diminuiu durante todo o ciclo de cultivo. A existência de um sumideiro grande (a raiz) unida a uma grande disponibilidade de substrato, permitiu que a estimulação da taxa fotosintética pelo [CO2] elevado continuasse no tempo. Foi avaliado o efeito de [CO2] muito altos sobre a fotosíntese de plantas silvestres que crescem ao redor de emanações naturais, descobrindo-se que tais concentrações não só não inibem a taxa fotosintética, senão que a promovem muito além dos controles.