Biomarcadores hematológicos del sapo común Bufo (Rhinella) arenarum en ecosistemas alterados de la provincia de Córdoba / Hematologic biomarkers of the common toad Bufo arenarum in altered ecosystem of Córdoba province

El uso de biomarcadores hematológicos es importante para poder evaluar riesgos ambientales y la exposición a sustancias químicas potencialmente tóxicas y así poder desarrollar medidas que sirvan como señales de alarma temprana en ambientes contaminados. El objetivo de este trabajo fue determinar biomarcadores hematológicos en el sapo común Bufo (Rhinella) arenarum que proporcionen información para el diagnóstico de la salud de distintos ecosistemas de la provincia de Córdoba. Un total de 93 individuos adultos de Bufo (Rhinella) arenarum fueron capturados en cuatro sitios de la provincia de Córdoba y distintas variables ambientales del agua fueron medidas en cada sitio. Las muestras de sangre se obtuvieron mediante la punción de la vena angularis. Luego fueron teñidas con la tinción diferencial May Grünwald-Giemsa y fueron observadas mediante microscopio Zeiss Primo Star iLED para realizar el conteo de glóbulos blancos. Las variables ambientales del agua mostraron diferencias estadísticamente significativas entre las poblaciones, se registraron valores menores en “Alpa Corral”, mientras que “Villa Dálcar” presentó los mayores valores para dichas variables. No se encontraron diferencias estadísticamente significativas entre localidades en el recuento de glóbulos blancos (cada 1000 eritrocitos); sin embargo, en “Villa Dálcar” se registraron los ejemplares con mayor cantidad de leucocitos en el conteo de glóbulos blancos (425,48 ± 481,34). Mientras que para la fórmula leucocitaria, los linfocitos fueron las células más abundantes para los ejemplares de todos los sitios; encontrándose diferencias estadísticamente significativas entre cada localidad y “Alpa Corral”. Los ejemplares de “Villa Dálcar” presentaron diferencias estadísticamente significativas en relación a los monocitos y basófilos, registrándose altos valores para ambas células en los ejemplares de este sitio; mientras que en los ejemplares de “Cultivo” las diferencias se encontraron en eosinófilos y la relación neutrófilos/linfocitos. Estos resultados revelaron que de las cuatro localidades analizadas, “Villa Dálcar” representa la más afectada por las actividades antrópicas. Estas variaciones registradas en los parámetros hematológicos podrían considerarse evidencia de la posible presencia de agentes contaminantes en el lago “Villa Dálcar”.

Hematologic biomarkers are important to assess the environmental and health risks of exposure to potentially toxic chemicals, and for developing measures that serve as early warning signals in polluted areas. The aim of this study was to determine hematologic biomarkers of common toad Bufo (Rhinella) arenarum that provide information for the diagnosis of the health of ecosystems of Córdoba province. A total of 93 adult individuals of Bufo (Rhinella) arenarum were collected from four sites of Córdoba Province and hydric environmental variables were measured on each site. Blood samples were obtained by angularis vein puncture, smears of fresh blood were stained with May Grünwald-Giemsa and observed by using a microscope Carl Zeiss Primo Star (Pack 5), for white blood cell count. Water environmental variables showed statistically significant differences among localities, with the lowest values in “Alpa Corral” and highest values in “Villa Dálcar”. There were no statistically significant differences among individuals of different sites for white blood cells count. However, individuals from "Villa Dálcar" showed the highest number of white cells (425.48 ± 481.34). In the leukocyte formula, lymphocytes were most abundant white blood cells in individuals from all study sites. The blood parameters of toads from all sites showed statistically significant differences with individuals from “Alpa Corral”. "Villa Dálcar" Individuals exhibited significant differences in relation to monocytes and basophils with the highest values for both cell types. “Cultivo” locality showed significant differences for eosinophils and for the relationship neutrophils/ lymphocytes. These results revealed that “Villa Dálcar” represent the site most affected by human activities. In particular, changes recorded in hematological parameters could be giving evidence of the possible presence of contaminants in Lake “Villa Dálcar”.

Demografía poblacional de Rhinella arenarum (Anura: Bufonidae) y Physalaemus biligonigerus (Anura: Leiuperidae) en agroecosistemas de la provincia de Córdoba, Argentina / Population demography in Rhinella arenarum (Anura: Bufonidae) and Physalaemus biligonigerus (Anura: Leiuperidae) in agroecosystems in the province of Córdoba, Argentina

The advancing agricultural frontier has led to an important loss of natural habitats, with significant consequences for biodiversity. The demography for two species of anurans, Physalaemus biligonigerus and Rhinella arenarum, both associated with agricultural systems in the central region of the Córdoba Province, were analyzed and compared in this study. Four sites were sampled: three agroecosystems with different alteration degrees (C1, C2 and SM1) and a fourth site not cultivated (SM2). The sampling was conducted during two reproductive periods, from September 2008 to April 2009 and September 2009 to April 2010. Individuals were captured using live pitfall traps for the metamorphic, juveniles and adults; and visual encounter survey, for the capture of eggs and larvae. With the abundance data, the survival for each age class was estimated using the KNM method (Kiritani Nakasuki Manly). With survival rates and fertility population, Leslie matrices were elaborated to obtain a quantitative projection of the population size. Altered environments showed lower eggs and larvae survival. Population pro-jections were favorable in the site SM2 and were less favorable and a tendency to extinction, in sites dominated by crops. This study showed that the agroecosystems of this region are possibly inhospitable environments for reproduction and survival of the species studied. The aquatic stages in the life cycle of both species would be the more affected, since water bodies deterioration is present or may occur in those areas. We can recognize species-specific effects of agricultural ecosystems; P. biligonigerus was the most affected species, possibly because of their life histories and habitat requirements. We suggested that environmental degradation caused by the cropland in the central region of Argentina would impact on the demographics of the anuran populations in the area.

El crecimiento de la agricultura produce pérdida del hábitat natural, con consecuencias para la biodiversidad de los anfibios. Se analizó la demografía poblacional de dos especies de anfibios en agroecosistemas de la provincia de Córdoba, Rhinella arenarum y Physalaemus biligonigerus. Los muestreos se realizaron en tres agroecosistemas y en un cuarto sitio, no afectado por cultivos. Los individuos se capturaron utilizando trampas de caída viva y relevamientos visuales. Con los datos de abundancia se estimó la supervivencia para cada clase de edad y con las tasas de supervivencia y fertilidad poblacional, se construyeron matrices de Leslie para obtener una proyección cuantitativa del tamaño poblacional. Las proyecciones poblacionales fueron favorables para el sitio SM2 y desfavorables para los sitios cultivados. Los agroecosistemas son ambientes posiblemente inhóspitos para la reproducción y supervivencia de las especies estudiadas. Particularmente se ven afectados aquellos estadios acuáticos del ciclo de vida de estas especies, debido al deterioro de los cuerpos de agua en estas áreas. Se reconocieron efectos especie-específicos de los ecosistemas agrícolas, ya que P. biligonigerus resultó la especie más afectada. Se sugiere un deterioro ambiental producido por cultivos extensivos de soja, lo que impactaría la demografía de las poblaciones de anuros.

Ecología trófica en larvas de Rhinella arenarum (Anura: Bufonidae) en agroecosistemas y sus posibles implicaciones para la conservación / Trophic ecology in tadpoles of Rhinella arenarum (Anura: Bufonidae) in agroecosystems and their possible implications for conservation

Trophic ecology in tadpoles of Rhinella arenarum (Anura: Bufonidae) in agroecosystems and their possible implications for conservation. The progress of the agriculture border has led an important loss of natural habitats, with significant consequences for biodiversity. In this sense, the studies in anuran amphibian tadpoles inhabiting these environments are relevant, because the larval stage is a phase of population regulation. The aim of this study was to analyze the body condition and diet in Rhinella arenarum, tadpoles, an anuran species widely distributed in South America and that inhabit agroecosystems. Three sites were sampled, two agroecosystems with different alteration degrees (C1 and C2) and an uncultured (SM) third place. The captured tadpoles were anesthetized, fixed and preserved in formaldehyde (10%). Subsequently, body measurements were made and the complete intestine was removed and analyzed for food items under a binocular microscope. The diet in R. arenarum tadpoles has a dominance of algae Bacillariophyceae, followed by Cyanophyceae. In particular, the class Bacillariophyceae, due to the presence of the genus Navicula, Nitzschia, Gomphonema and Hantzschia, was important in the diet of the anurans in those agroecosystems. Class Cyanophyceae, mainly represented by genus Oscillatoria and Euglenophyceae represented by Euglena and Strombomonas, were predominant in the diet of the anurans in SM. Some differences in the total items consumed by tadpole were observed between the studied sites. Tadpoles that inhabit the modified sites (C1 and C2) recorded a significantly smaller amount of food. Moreover, the tadpoles that inhabit these sites showed a lower body condition. The presence of certain algae associated with eutrophic environments, could indicate some pollution in agroecosystems (C1 and C2). Food resources would be lesser in places with strong agricultural activity, possibly with a greater degree of eutrophication. A smaller food amount could have consequences at population level for the short and long time terms, because of its impact on individual growth. Larval diet is suggested as a potential bioindicator of environmental health for these areas. Rev. Biol. Trop. 60 (2): 771-779. Epub 2012 June 01.

El crecimiento de la agricultura produce perdida del habitat natural, con consecuencias para la biodiversidad de los anfibios. Se analizo la dieta y condición corporal de larvas de anuros de Rhinella arenarum que habitan agroecosistemas. Los muestreos fueron realizados en dos agroecosistemas y en un tercer sitio, no afectado por cultivos. Las larvas capturadas fueron anestesiadas, fijadas y preservadas en formaldehido, se realizaron medidas corporales y se analizo la dieta de las larvas. Se registro diversidad en la oferta alimenticia proporcionada por los distintos ambientes. La dieta tuvo una predominancia de algas Bacillarophyceae, seguidas por Cyanophyceae. Particularmente, los géneros Navicula, Nitzschia, Hantzschia y Gomphonema (clase Bacillarophyceae), fueron importantes en los agroecosistemas. Los géneros Osillatoria, Euglena y Strombomonas (clases Cyanophyceae y Euglenophyceae), predominaron en la dieta de las larvas de anuros en el sitio menos alterado. Las larvas de los sitios más alterados consumen menor cantidad de alimento y registran una menor condición corporal. La presencia de determinadas algas, indicarían un mayor grado de contaminación en los agroecosistemas. La eutrofización de lagunas podría alterar la disponibilidad de alimento para larvas de anuros que podría tener consecuencias poblacionales negativas. Se sugiere el análisis de dietas larvarias como un potencial bioindicador de salubridad ambiental.

Thermal parameters changes in males of Rhinella arenarum (Anura:Bufonidae) related to reproductive periods

The regulation of body temperature in ectotherms has a major impact in their physiological and behavioral processes. Observing changes in thermal parameters related with reproduction allows us to better understand how Rhinella arenarum optimizes a thermal resource. The aim of this study was to compare the thermal parameters of this species between breeding and non-breeding periods. In the field, we recorded the body temperature from captured animals, the air temperature, and the temperature of the substrate. In the laboratory, we measured the temperature R. arenarum selected on a thermal gradient and the critical extreme temperatures. The results of our study show variations in thermal parameters between the two situations studied. This species makes efficient use of thermal resources during the breeding period by basking to significantly increase body temperature. Because calling is energetically costly for males, this behavior results in increased efficiency to callers during the breeding period. Rev. Biol. Trop. 59 (1): 347-353. Epub 2011 March 01.

La regulación de la temperatura en ectotérmos tiene gran importancia en los procesos fisiológicos y comportamentales. Los cambios en los parámetros térmicos relacionados con la reproducción nos permiten entender de qué manera Rhinella arenarum optimiza el recurso térmico. El objetivo del presente trabajo fue comparar los parámetros térmicos de la especie entre el periodo reproductivo y no reproductivo. En el campo se registraron la temperatura corporal de los animales capturados, la temperatura del aire y del sustrato. Además, en laboratorio se registro la temperatura selecta en un gradiente térmico. Como así también las temperaturas criticas máxima y mínima. Los resultados de nuestro estudio muestran variaciones de los parámetros térmicos entre ambas situaciones estudiadas. Aparentemente esta especie hace un uso eficiente del recurso térmico durante el periodo reproductivo ya que el basking le permite aumentar considerablemente la temperatura corporal, probablemente este comportamiento esté relacionado con el aumento de la eficacia para emitir el canto durante el periodo de reproducción ya que el mismo es energéticamente costoso para los machos.

Water-balance response of Rhinella arenarum (Hensel, 1867) tadpoles to graduated increase in environmental osmolarity / Resposta de Rhinella arenarum larvae a meio electrolítico e não electrolítico com aumento escalonado da osmolaridade

The water balance and the upper limit of osmotic tolerance of premetamorphic Rhinella arenarum larvae (Gosner's stage 26) was evaluated after semistatic incubation in electrolyte (NaCl) and non-electrolyte (mannitol) media following a protocol of progressively increased osmotic pressure. Wet and dry weights were measured to calculate the water content as a derived variable indicative of the hydric balance. Statistical analysis was performed using univariate and integrated multivariate analysis. Tadpoles survived in electrolyte and non-electrolyte solutions up to 200 mOsm. The discriminant function was the best tool to describe the responses of the animals to external environmental stress under experimental conditions. The results were compared with those obtained in previous studies using a protocol of acute exposure to the same media used in this study. It was concluded that a) multivariate analysis is an appropriate approach to describe the responses of tadpoles to changes in the environmental physicochemical parameters, and b) progressive and acute acclimation to the experimental solutions induced similar responses.

Avaliou-se o balanço hídrico e o limite superior de tolerância osmótica em larvas pré-metamórficas do Rhinella arenarum (etapa 26 de Gosner) sob condições de incubação semiestáticas, num meio eletrolítico (NaCl) e não eletrolítico (manitol), seguindo um protocolo de aumento progressivo da pressão osmótica do meio. A quantificação das respostas se efetuou por meio da medição dos valores de peso úmido e seco e do cálculo, a partir destes, do conteúdo de água, como variável derivada indicativa do equilíbrio hídrico. A análise estatística foi realizada usando análise univariada e multivariada. As larvas conseguiram sobreviver em soluções eletrolíticas e não eletrolíticas até 200 mOsm. A função discriminante foi a melhor ferramenta para descrever as respostas dos animais ao estresse osmótico ambiental. Os resultados foram comparados com os obtidos em estudos anteriores, usando um protocolo de exposição aguda aos mesmos meios de incubação usados neste estudo. Concluiu-se: a) a análise multivariada é a aproximação adequada para descrever as respostas das larvas às mudanças nos parâmetros físico-químicos do seu meio; e b) tanto a aclimatação progressiva, como as exposições agudas às soluções experimentais induziram as respostas semelhantes.