Identification of suitable internal control genes for expression studies in Coffea arabica under different experimental conditions

Background: Quantitative data from gene expression experiments are often normalized by transcription levels of reference or housekeeping genes. An inherent assumption for their use is that the expression of these genes is highly uniform in living organisms during various phases of development, in different cell types and under diverse environmental conditions. To date, the validation of reference genes in plants has received very little attention and suitable reference genes have not been defined for a great number of crop species including Coffea arabica. The aim of the research reported herein was to compare the relative expression of a set of potential reference genes across different types of tissue/organ samples of coffee. We also validated the expression profiles of the selected reference genes at various stages of development and under a specific biotic stress.Results: The expression levels of five frequently used housekeeping genes (reference genes), namely alcohol dehydrogenase (adh), 14-3-3, polyubiquitin (poly), beta-actin (actin) and glyceraldehyde-3-phosphate dehydrogenase (gapdh) was assessed by quantitative real-time RT-PCR over a set of five tissue/organ samples (root, stem, leaf, flower, and fruits) of Coffea arabica plants. In addition to these commonly used internal controls, three other genes encoding a cysteine proteinase (cys), a caffeine synthase (ccs) and the 60S ribosomal protein L7 (rpl7) were also tested. Their stability and suitability as reference genes were validated by geNorm, NormFinder and BestKeeper programs. The obtained results revealed significantly variable expression levels of all reference genes analyzed, with the exception of gapdh, which showed no significant changes in expression among the investigated experimental conditions.Conclusion: Our data suggests that the expression of housekeeping genes is not completely stable in coffee. Based on our results, gapdh, followed by 14-3-3 and rpl7 were found to be homogeneously expressed and are therefore adequate for normalization purposes, showing equivalent transcript levels in different tissue/ organ samples. Gapdh is therefore the recommended reference gene for measuring gene expression in Coffea arabica. Its use will enable more accurate and reliable normalization of tissue/organ-specific gene expression studies in this important cherry crop plant.

Infection levels of Austrodiplostomum compactum (Digenea, Diplostomidae) metacercariae in Plagioscion squamosissimus (Teleostei, Sciaenidae) from the Nova Avanhandava reservoir, São Paulo State, Brazil

Conselho Nacional de Desenvolvimento Científico e Tecnológico (CNPq)

Ecological aspects of hermit crabs (Crustacea, Anomura, Paguroidea) off the northern coast of São Paulo State, Brazil

The occurrence of species of hermit crabs and their ecological distribution in soft bottoms off Ubatuba, São Paulo, Brazil were analyzed. To better understand the distribution of the species in relation to environmental factors, the similarity and species-diversity indexes were calculated. Paguroideans were sampled monthly from January through December 2000. The trawls were made with two otter-trawl nets at 13 different sites, at depths of 2-40 m. Water temperature, salinity, sediment texture, and organic matter content were measured. Gastropod shells occupied by hermit crabs were also assessed. A total of 1,238 specimens was collected, belonging to the families Diogenidae and Paguridae, comprising seven genera and thirteen species. The most abundant hermit crab species were Dardanus insignis (761 specimens) and Loxopagurus loxochelis (351 specimens). Phimochirus holthuisi is newly reported from the São Paulo coast. The highest diversity index was found for the shallower sites near rocky shores. The results of the grouping analysis for sites and species indicated three distinct groups for sites, and four groups for species. This suggests that the occurrence of these anomurans is associated with the environmental and biotic factors analyzed.

Fishes of the Atlantic Rainforest of Boraceia: testimonies of the Quaternary fault reactivation within a Neoproterozoic tectonic province in Southeastern Brazil

The upper portion of the Rio Guaratuba, located in the Serra do Mar coastal range of Southeastern Brazil, shows biological and geological evidences of being captured and diverted away from its original course in the direction of the Parana River Basin to become a coastal river due to the Quaternary activity of NW-trending faults. Despite draining directly to the Atlantic Ocean, the upper portion of the Guaratuba still maintains an ichthyofauna that is typical of the adjoining Parana River Basin rather than the characteristic fish fauna of the Brazilian coastal drainages. The fish fauna of the upper Guaratuba is an evident testimony of the tectonic process that allowed the faunal interchange between the upland basins and the coastal drainages that probably has been taking place in Southeastern Brazil throughout the long geological history of the passive Brazilian continental margin.

Marine anomurans (Decapoda) from the non-consolidated sublittoral bottom at the southeastern coast of Brazil

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Forest fragment size and microhabitat effects on palm seed predation

The establishment of plant species depends crucially on where the seeds are deposited. However, since most studies have been conducted in continuous forests, not much is known about the effects of forest fragmentation on the maintenance of abiotic and biotic characteristics in microhabitats and their effects on seed survival. in this study, we evaluated the effects of forest fragmentation on the predation upon the seeds of the palm Syagrus romanzoffiana in three microhabitats (interior forest, forest edge and gaps) in eight fragments of semi-deciduous Atlantic forest ranging in size from 9.5 ha to 33,845 ha in southeastern Brazil. Specifically, we examined the influence of the microhabitat structure, fauna and fragment size on the pattern of seed predation. Fragments < 100 ha showed similar abiotic and biotic characteristics to those of the forest edge, with no seed predation in these areas. Forest fragments 230-380 ha in size did not present safe sites for S. romanzoffiana seed survival and showed high seed predation intensity in all microhabitats evaluated. In fragments larger than 1000 ha, the seed predation was lower, with abiotic and biotic differences among gaps, interior forests and forest edges. In these fragments, the survival of S. romanzoffiana seeds was related to squirrel abundance and interior forest maintenance. Based on these results, we concluded that there are no safe sites for S. romanzoffiana seed establishment in medium- and small-sized fragments as result of the biotic and abiotic pressure, respectively We suggest that on these forest fragments, management plans are needed for the establishment of S. romanzoffiana, such as interior forest improvement and development in small-sized sites in order to minimize the edge effects, and on medium-sized fragments, we suggest post-dispersal seed protection in order to avoid seed predation by vertebrates. our findings also stress the importance of assessing the influence of forest fragmentation on angiosperm reproductive biology as part of the effective planning for the management of fragmented areas. (c) 2006 Elsevier Ltd. All rights reserved.

Behavior of Ectatomma brunneum (Formicidae: Ectatomminae) Preying on Dipterans in Field Conditions

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Genotoxicity assessment of water soluble fractions of biodiesel and its diesel blends using the Salmonella assay and the in vitro MicroFlow (R) kit (Litron) assay

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Rhamnolipids and PHAs: Recent reports on Pseudomonas-derived molecules of increasing industrial interest

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Seed size variation in the palm Euterpe edulis and the effects of seed predators on germination and seedling survival

Intraspecific variation in seed size is common in wild plant populations and has important consequences for the reproductive success of individual plants. Multiple, often conflicting evolutionary forces mediated by biotic as well as abiotic agents may maintain such a variation. In this paper we assessed seed size variation in a population of the threatened, commercially important palm Euterpe edulis in southeast Brazil. We investigated (i) how this variation affects the probability of attack by vertebrate and invertebrate post-dispersal seed predators, and (ii) if seed size influences the outcome of seeds damaged by beetles in terms of seed germination and early survival of seedlings. Euterpe edulis seeds varied in diameter from 8.3 to 14.1 mm. Neither insects nor rodents selected the seeds they preyed upon based on seed size. Seed germination and total, shoot and root biomasses of one-year seedlings were significantly and positively affected by seed size. Root biomass and seedling survival were negatively affected by seed damage caused by a scolytid beetle (Coccotrypes palmarum) whose adults bore into seeds to consume part of the endosperm, but do not oviposit on them. Seed size had a marginally significant effect on seedling survival. Therefore, if any advantage is accrued by E. edulis individuals producing large seeds, this is because of greater seed germination success and seedling vigor. If this is so, even a relatively narrow range of variation in seed size as observed in the E. edulis population studied may translate into differential success of individual plants. (c) 2006 Elsevier SAS. All rights reserved.

The shared influence of phylogeny and ecology on the reproductive patterns of Myrteae (Myrtaceae)

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Leaf structure of the cerrado (Brazilian savanna) woody plants

Fundação de Amparo à Pesquisa do Estado de São Paulo (FAPESP)

Contrasting responses to induction cues by ants inhabiting Maieta guianensis (Melastomataceae)

As duas formigas que mais comumente se associam com a mirmecofita de sub-bosque Maieta guianensis (Melastomataceac) respondem de maneiras diferentes a simulac (o) over tilde es experimentais de herbivoria. Enquanto Pheidole minutula & induzida tanto por danos fisicos e extratos de tecido foliar, Crematogaster laevis 6 induzida apenas pelos danos fisicos. Sugerimos que tal variac (a) over tildeo interespecifica em respostas induzidas e comum e pode influenciar a qualidade das defesas contra herbivoros que as formigas oferecem as plantas que elas ocupam.

Interspecific variation in the defensive responses of obligate plant-ants: experimental tests and consequences for herbivory

The aggressive behavior of ants that protect plants from herbivores in exchange for rewards such as shelter or food is thought to be an important form of biotic defense against herbivory, particularly in tropical systems. To date, however, no one has compared the defensive responses of different ant taxa associated with the same plant species, and attempted to relate these differences to longer-term efficacy of ant defense. We used experimental cues associated with herbivory-physical damage and extracts of chemical volatiles from leaf tissue-to compare the aggressive responses of two ant species obligately associated with the Amazonian myrmecophyte Tococa bullifera (Melastomataceae). We also conducted a colony removal experiment to quantify the level of resistance from herbivores provided to plants by each ant species. Our experiments demonstrate that some cues eliciting a strong response from one ant species elicited no response by the other. For cues that do elicit responses, the magnitude of these responses can vary interspecifically. These patterns were consistent with the level of resistance provided from herbivores to plants. The colony removal experiment showed that both ant species defend plants from herbivores: however, herbivory was higher on plants colonized by the less aggressive ant species. Our results add to the growing body of literature indicating defensive ant responses are stimulated by cues associated with herbivory. However, they also suggest the local and regional variation in the composition of potential partner taxa could influence the ecology and evolution of defensive mutualisms in ways that have previously remained unexplored.

Adaptation of the reservoir fish assemblage index (RFAI) for assessing the Barra Bonita Reservoir (São Paulo, Brazil)

In order to evaluate the environmental condition of the Barra Bonita Reservoir, we adapted the Index of Biotic Integrity (IBI). We chose 24 sampling sites in which three types of habitats were sampled: the mouths of tributaries, the central reservoir and the lateral reservoir. Fish were caught in two seasons (dry and rainy) using 10 gillnet gangs, with meshes ranging from 3 to 12 cm between opposite knots, and funnel traps. Abiotic and biotic variables were measured. Due to the artificial nature of the reservoir, the term biotic integrity was considered inappropriate and the term RFAI (Reservoir Fish Assemblage Index) was adopted. Twelve metrics from sixteen possibles were selected using the Pearson correlation coefficient. The reference conditions were set up based on the criteria of the best condition observed. For each metric, a score of 1, 3 or 5 was assigned as it strongly departs (1), slightly departs (3) or approaches (5) the reference condition. The index value is the sum of the metrics partial scores and ranges from 12 to 60. To evaluate the importance of the unit of measurement of the metric, the index was calculated in fish number (RFAI(N)) and in weight (RFAI(w)). The correlation between RFAIN and RFAIw was very high (r = 0.90, n = 46) indicating that the unit of measurement does not influence the final result of the index. Most of the sampling sites were classified in the 'reasonable' RFAI category. Only the central sites were classified as 'poor'. To validate the RFAI, another index, the Habitat Quality Index (HQI), was built starting from the physiochemical and habitat variables collected. The correlation of the RFAI with the HQI was highly significant (RFAIN, r = 0.37; RFAI(w), r = 0.47; n = 46), indicating that they respond in the same way to environmental degradation. The HQI metrics which most affect RFAI were depth, surrounding landscape and macrophyte presence/absence. Copyright C) 2007 John Wiley & Sons, Ltd.