Clonal growth and serial propagation of rat esophageal epithelial cells

The clonal growth and serial propagation of rat esophageal epithelial cells in low serum-containing medium has been achieved without feeder layers or conditioned medium. To date, a total of four lines have been developed and maintained for as many as 40 passages in culture. Growth of the cells was possible only after modifying the culture medium (PFMR-4) by reducing the calcium concentration from 1 to 0.1 mM, and by adding low levels of dialyzed fetal bovine serum and seven growth factors; i.e. epidermal growth factor, hydrocortisone, ethanolamine, phosphoethanolamine, insulin, transferrin, and cholera toxin. Cell lines have been developed from both explant outgrowths and enzyme dissociated esophagi. The epithelial nature of the cells was confirmed by electron microscopy and immunological methods. Clonal growth studies revealed that optimal cell growth occurred in medium containing 2.4% dialyzed fetal bovine serum and 0.1 mM calcium. Calcium levels of 0.3 mM or higher caused the cells to stratify and undergo terminal differentiation. Coating the culture dishes with collagen, or a combination of collagen, fibronectin, and bovine serum albumin, increased both the cell growth rate and the colony forming efficiency. The successful long term culture of rat esophageal epithelial cells permits their use as models in studies concerned with esophageal differentiation and carcinogenesis.     

Seasonal intake responses in the nectar-feeding bat Glossophaga soricina

Food intake in nectar-feeding animals is affected by food quality, their energetic demands, and the environmental conditions they face. These animals increase their food intake in response to a decrease in food quality, a behavior named “intake response”. However, their capacity to achieve compensatory feeding, in which they maintain a constant flux of energy, could be constrained by physiological processes. Here we evaluated how both a seasonal change in environmental conditions and physiological constraints affected the food ingestion in the bat Glossophaga soricina. We measured food intake rate during both the wet/warm and dry/cool seasons at sucrose solutions ranging from 146 to 1,022 mmol L⁻¹. We expected that food intake and metabolic demands would be greater during the dry/cool season. Bats ingested ~20% more food in the dry/cool than in the wet/warm season. Regardless of season, bats were unable to achieve a constant flux of energy when facing the different sugar concentrations that we used in our experiments. This suggests that the rate of food intake is physiologically constrained in G. soricina. Using the digestive capacity of bats we modeled their food intake. The analytic model we used predicts that digestive limitations to ingest energy should have an important effect on the ecology of this species.     

Movement of soil-applied imidacloprid and thiamethoxam into nectar and pollen of squash (Cucurbita pepo)

There has been recent interest in the threat to bees posed by the use of systemic insecticides. One concern is that systemic insecticides may translocate from the soil into pollen and nectar of plants, where they would be ingested by pollinators. This paper reports on the movement of two such systemic neonicotinoid insecticides, imidacloprid and thiamethoxam, into the pollen and nectar of flowers of squash (Cucurbita pepo cultivars "Multipik," "Sunray" and "Bush Delicata") when applied to soil by two methods: (1) sprayed into soil before seeding, or (2) applied through drip irrigation in a single treatment after transplant. All insecticide treatments were within labeled rates for these compounds. Pollen and nectar samples were analyzed using a standard extraction method widely used for pesticides (QuEChERS) and liquid chromatography mass spectrometric analysis. The concentrations found in nectar, 10 ± 3 ppb (mean ± s.d) for imidacloprid and 11 ± 6 ppb for thiamethoxam, are higher than concentrations of neonicotinoid insecticides in nectar of canola and sunflower grown from treated seed, and similar to those found in a recent study of neonicotinoids applied to pumpkins at transplant and through drip irrigation. The concentrations in pollen, 14 ± 8 ppb for imidacloprid and 12 ± 9 ppb for thiamethoxam, are higher than those found for seed treatments in most studies, but at the low end of the range found in the pumpkin study. Our concentrations fall into the range being investigated for sublethal effects on honey bees and bumble bees.     

Massive Destruction of Symphonia globulifera (Clusiaceae) Flowers by Central American Spider Monkeys (Ateles geoffroyi)1

Although more than 39 primate species have been described as nectar feeders, no studies have documented the negative affect this behavior may have on the reproductive success of the plants they consume. Here we report, for the first time, massive flower destruction of the tropical tree Symphonia globulifera by the frugivorous spider monkey, Ateles geoffroyi, and document the detrimental effect this behavior has on fruit set. Foraging behavior was collected from one troop of A. geoffroyi during 460 contact hours from June 1999 to May 2000 in the tropical humid forest at the Refugio de Vida Silvestre Punta Rio Claro (8°39′N, 83°44′E) in the Osa Peninsula in Southwestern Costa Rica. Detailed phenological data were collected from ten S. globulifera trees that the monkeys fed upon and ten trees that were outside of their home range, but in the Refugio. From July to September 1999 S. globulifera was the most important species consumed, representing from 86 to 100 percent of the total feeding time each month. Monkey foraging on S. globulifera flowers was destructive, detaching the flowers completely from the branches in 80 percent of the foraging bouts, and leaving flowers without petals and with damaged reproductive parts in 20 percent of the bouts. None of the ten trees where the monkeys were foraging set fruit, but seven of ten trees outside of their home range set fruit in November and December 1999. These results suggest that some primates may play an important role in floral herbivory in tropical forests and the subsequent reduction in reproductive success. Future studies should focus on evaluating the long‐term effect this foraging behavior may have on the population structure of this species and evaluate the effect of primate floral herbivory on the reproductive success of other plant species.     

Isolation and characterization of epithelial cells from bovine pancreatic duct

Epithelial cells derived from bovine pancreatic duct have been grown continuously in culture for 30 weeks (approximately 90 doublings of the cell population). The cells were grown in Eagle's minimal essential medium supplemented with 10% heat-inactivated fetal bovine serum, 2 mM glutamine, 0.1 mM nonessential amino acids, and antibiotics. In confluent cultures, the cells are multilayered and form circular structures. When tested at various passages, the cells neither formed colonies in soft agar nor produced tumors after inoculation into athymic, nude mice. Hydrocortisone (1 and 5 microgram per ml) and insulin (1,5 and 10 microgram per ml) had no effect on the growth of the cells. beta-Retinyl acetate inhibited growth rate and cell yield at a concentration of 5 microgram per ml but was not growth-inhibitory at lower concentrations. By electron microscopy the cells have numerous mitochondria, Golgi and microvilli. Mucous droplets were observed in a small proportion of the cells. Desmosome-like structures and occluding junctions were observed more frequently between cells that had been transferred as aggregates than between cells transferred as single cells. Cytochemical studies indicated that some cells produce PAS positive granules that were not removed after treatment of the cultures with diastase. Eleven cell clones were isolated from the mass culture. The growth rates of the clones are different as well as the period of time in which the clones can be propagated in vitro.     

Sleeping Sites and Latrines of Spider Monkeys in Continuous and Fragmented Rainforests: Implications for Seed Dispersal and Forest Regeneration

Spider monkeys (Ateles geoffroyi) use sites composed of one or more trees for sleeping (sleeping sites and sleeping trees, respectively). Beneath these sites/trees they deposit copious amounts of dung in latrines. This behavior results in a clumped deposition pattern of seeds and nutrients that directly impacts the regeneration of tropical forests. Therefore, information on the density and spatial distribution of sleeping sites and latrines, and the characteristics (i.e., composition and structure) of sleeping trees are needed to improve our understanding of the ecological significance of spider monkeys in influencing forest composition. Moreover, since primate populations are increasingly forced to inhabit fragmented landscapes, it is important to assess if these characteristics differ between continuous and fragmented forests. We assessed this novel information from eight independent spider monkey communities in the Lacandona rainforest, Mexico: four continuous forest sites and four forest fragments. Both the density of sleeping sites and latrines did not differ between forest conditions. Latrines were uniformly distributed across sleeping sites, but the spatial distribution of sleeping sites within the areas was highly variable, being particularly clumped in forest fragments. In fact, the average inter-latrine distances were almost double in continuous forest than in fragments. Latrines were located beneath only a few tree species, and these trees were larger in diameter in continuous than fragmented forests. Because latrines may represent hotspots of seedling recruitment, our results have important ecological and conservation implications. The variation in the spatial distribution of sleeping sites across the forest indicates that spider monkeys likely create a complex seed deposition pattern in space and time. However, the use of a very few tree species for sleeping could contribute to the establishment of specific vegetation associations typical of the southeastern Mexican rainforest, such as Terminalia-Dialium, and Brosimum-Dialium.