Ability of Salmonella spp. to Produce Biofilm Is Dependent on Temperature and Surface Material

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

Predicting embryo presence and viability

Pregnancy establishment, followed by birth of live offspring, is essential to all mammals. The biological processes leading up to pregnancy establishment, maintenance, and birth are complex and dependent on the coordinated timing of a series of events at the molecular, cellular, and physiological level. The ability to ovulate a competent oocyte, which is capable of undergoing fertilization, is only the initial step in achieving a successful pregnancy. Once fertilization has occurred and early embryonic development is initiated, early pregnancy detection is critical to provide proper prenatal care (humans) or appropriate management (domestic livestock). However, the simple presence of an embryo, early in gestation, does not guarantee the birth of a live offspring. Pregnancy loss (embryonic mortality, spontaneous abortions, etc.) has been well documented in all mammals, especially in humans and domestic livestock species, and is a major cause of reproductive loss. It has been estimated that only about 25-30 % of all fertilized oocytes in humans result in birth of a live offspring; however, identifying the embryos that will not survive to parturition has not been an easy task. Therefore, investigators have focused the identification of products in maternal circulation that permit the detection of an embryo and assessment of its well-being. This review will focus on the advances in predicting embryonic presence and viability, in vivo.

Effects of UV-B radiation on the antagonistic ability of Clonostachys rosea to Botrytis cinerea on strawberry leaf disc

Human activities are altering the concentrations of ozone in the troposphere and hence in the incidence of ultraviolet-B (UV-B) on Earth’s surface. Although representing only five percent of UV-B radiation striking the Earth's surface, this radiation has the potential to cause effects on biologically active molecules. Sensitivity to UV-B radiation is one of the limitations of biological control of plant pathogens in the field. The objectives of this work were to evaluate the effects of UV-B on several isolates of Clonostachys rosea, and the ability of an isolate of C. rosea, previously selected for its tolerant to UV-B radiation, to control Botrytis cinerea on strawberry leaves in controlled conditions (strawberry leaf discs). The germination of C. rosea conidia was inversely proportional to the irradiance. The most tolerant strain (LQC62) had relative germination of about 60% after irradiation of 4.2kJ/m2, and this strain was selected to be used in the subsequent studies. The data showed that even with exposure to UV-B radiation, C. rosea LQC62 controlled the pathogen. Conidial concentrations of strain LQC62 above 105 conidia/ml showed higher tolerance to UV-B radiation and increased ability to control more than 75% of the B. cinerea even with exposure to radiation. According to our results, in addition to showing less growth under UV-B, conidia of C. rosea had lower antagonistic ability. Further studies are needed to observe the tolerance of B. cinerea conidia to UV-B radiation and thereby prove that an environment with increased UV-B radiation may be favoring the pathogen due to a lower ability of C. rosea to control the pathogen in conditions of increased UV-B.

A Novel Method of Evaluating Ureteropelvic Junction Obstruction: Dynamic Near Infrared Fluorescence Imaging Compared to Standard Modalities to Assess Urinary Obstruction in a Swine Model

Purpose: Dynamic near infrared fluorescence imaging of the urinary tract provides a promising way to diagnose ureteropelvic junction obstruction. Initial studies demonstrated the ability to visualize urine flow and peristalsis in great detail. We analyzed the efficacy of near infrared imaging in evaluating ureteropelvic junction obstruction, renal involvement and the anatomical detail provided compared to conventional imaging modalities. Materials and Methods: Ten swine underwent partial or complete unilateral ureteral obstruction. Groups were survived for the short or the long term. Imaging was performed with mercaptoacetyltriglycine diuretic renogram, magnetic resonance urogram, excretory urogram, ultrasound and near infrared imaging. Scoring systems for ureteropelvic junction obstruction were developed for magnetic resonance urogram and near infrared imaging. Physicians and medical students graded ureteropelvic junction obstruction based on magnetic resonance urogram and near infrared imaging results. Results: Markers of vascular and urinary dynamics were quantitatively consistent among control renal units. The same markers were abnormal in obstructed renal units with significantly different times of renal phase peak, start of pelvic phase and start of renal uptake. Such parameters were consistent with those obtained with mercaptoacetyltriglycine diuretic renography. Near infrared imaging provided live imaging of urinary flow, which was helpful in identifying the area of obstruction for surgical planning. Physicians and medical students categorized the degree of obstruction appropriately for fluorescence imaging and magnetic resonance urogram. Conclusions: Near infrared imaging offers a feasible way to obtain live, dynamic images of urine flow and ureteral peristalsis. Qualitative and quantitative parameters were comparable to those of conventional imaging. Findings support fluorescence imaging as an accurate, easy to use method of diagnosing ureteropelvic junction obstruction.

Functional dna nanostructures for in vitro biosensing in live cells

DNA is a fascinating biomolecule that is well known for its genetic role in living systems. The emerging area of DNA nanotechnology provides an alternative view that exploits unparallel self-assembly ability of DNA molecules for material use of DNA. Although many reports exist on the results of DNA self-assembling systems, still few of them focus on the in vitro study about the function of such DNA nanostructures in live cells. Due to this, there are still a limited research about the in vitro functionality of such designs. To address an aspect of this issue, we have designed, synthesized and characterized two multifunctional fluorescencent nanobiosensors by DNA self-assembling. Each structure was designed and implemented to be introduced in live cells in order to give information on their functioning in real-time. Computational tools were used in order to design a graphic model of two new DNA motifs and also to obtain the specific sequences to all the ssDNA molecules. By thermal self-assembly techniques we have successfully synthesized the structure and corroborate their formation by the PAGE technique. In addition, we have established the conditions to characterize their structural conformation change when they perform their sensor response. The sensing behavior was also accomplished by fluorescence spectroscopy techniques; FRET evaluation and fluorescence microscopy imaging. Providing the evidence about their adequate sensing performance outside and inside the cells detected in real-time. In a preliminary evaluation we have tried to show the in vitro functionality of our structures in different cancer cell lines with the ability to perform local sensing responses. Our findings suggest that DNA sensor nanostructures could serve as a platform to exploit further therapeutic achievements in live cells.

Micropollutants affect the ability of phytoplankton communities to track environmental changes

Human activities strongly influence environmental processes, and while human domination increases, biodiversity progressively declines in ecosystems worldwide. High genetic and phenotypic variability ensures functionality and stability of ecosystem processes through time and increases the resilience and the adaptive capacity of populations and communities, while a reduction in functional diversity leads to a decrease in the ability to respond in a changing environment. Pollution is becoming one of the major threats in aquatic ecosystem, and pharmaceutical and personal care products (PPCPs) in particular are a relatively new group of environmental contaminants suspected to have adverse effects on aquatic organisms. There is still a lake of knowledge on the responses of communities to complex chemical mixtures in the environment. We used an individual-trait-based approach to assess the response of a phytoplankton community in a scenario of combined pollution and environmental change (steady increasing in temperature). We manipulated individual-level trait diversity directly (by filtering out size classes) and indirectly (through exposure to PPCPs mixture), and studied how reduction in trait-diversity affected community structure, production of biomass and the ability of the community to track a changing environment. We found that exposure to PPCPs slows down the ability of the community to respond to an increasing temperature. Our study also highlights how physiological responses (induced by PPCPs exposure) are important for ecosystem processes: although from an ecological point of view experimental communities converged to a similar structure, they were functionally different.

The Use of Human and Conspecific Gestures By Capuchin Monkeys to Solve an Object-Choice Task

Most primates live in highly complex social systems, and therefore have evolved similarly complex methods of communicating with each other. One type of communication is the use of manual gestures, which are only found in primates. No substantial evidence exists indicating that monkeys use communicative gestures in the wild. However, monkeys may demonstrate the ability to learn and/or use gestures in certain experimental paradigms since they¿ve been shown to use other visual cues such as gaze. The purpose of this study was to investigate if ten brown capuchin monkeys (Cebus apella) were able to use gestural cues from monkeys and a pointing cue from a human to obtain a hidden reward. They were then tested to determine if they could transfer this skill from monkeys to humans and from humans to monkeys. One group of monkeys was trained and tested using a conspecific as the cue giver, and was then tested with a human cue-giver. The second group of monkeys began training and testing with a human cue giver, and was then tested with a monkey cue giver. I found that two monkeys were able to use gestural cues from conspecifics (e.g., reaching) to obtain a hidden reward and then transfer this ability to a pointing cue from a human. Four monkeys learned to use the human pointing cue first, and then transferred this ability to use the gestural cues from conspecifics to obtain a hidden reward. However, the number of trials it took for each monkey to transfer the ability varied considerably. Some subjects spontaneously transferred in the minimum number of trials needed to reach my criteria for successfully obtaining hidden rewards (N = 40 trials), while others needed a large number of trials to do so (e.g. N = 190 trials). Two subjects did not perform successfully in any of the conditions in which they were tested. One subject successfully used the human pointing cue and a human pointing plus vocalization cue, but did not learn the conspecific cue. One subject learned to use the conspecific cue but not the human pointing cue. This was the first study to test if brown capuchin monkeys could use gestural cues from conspecifics to solve an object choice task. The study was also the first to test if capuchins could transfer this skill from monkeys to humans and from humans to monkeys. Results showed that capuchin monkeys were able to flexibly use communicative gestures when they were both unintentionally given by a conspecific and intentionally given by a human to indicate a source of food.

Biological safety concepts of genetically modified live bacterial vaccines

Live vaccines possess the advantage of having access to induce cell-mediated and antibody-mediated immunity; thus in certain cases they are able to prevent infection, and not only disease. Furthermore, live vaccines, particularly bacterial live vaccines, are relatively cheap to produce and easy to apply. Hence they are suitable to immunize large communities or herds. The induction of both cell-mediated immunity as well as antibody-mediated immunity, which is particularly beneficial in inducing mucosal immune responses, is obtained by the vaccine-strain's ability to colonize and multiply in the host without causing disease. For this reason, live vaccines require attenuation of virulence of the bacterium to which immunity must be induced. Traditionally attenuation was achieved simply by multiple passages of the microorganism on growth medium, in animals, eggs or cell cultures or by chemical or physical mutagenesis, which resulted in random mutations that lead to attenuation. In contrast, novel molecular methods enable the development of genetically modified organisms (GMOs) targeted to specific genes that are particularly suited to induce attenuation or to reduce undesirable effects in the tissue in which the vaccine strains can multiply and survive. Since live vaccine strains (attenuated by natural selection or genetic engineering) are potentially released into the environment by the vaccinees, safety issues concerning the medical as well as environmental aspects must be considered. These involve (i) changes in cell, tissue and host tropism, (ii) virulence of the carrier through the incorporation of foreign genes, (iii) reversion to virulence by acquisition of complementation genes, (iv) exchange of genetic information with other vaccine or wild-type strains of the carrier organism and (v) spread of undesired genes such as antibiotic resistance genes. Before live vaccines are applied, the safety issues must be thoroughly evaluated case-by-case. Safety assessment includes knowledge of the precise function and genetic location of the genes to be mutated, their genetic stability, potential reversion mechanisms, possible recombination events with dormant genes, gene transfer to other organisms as well as gene acquisition from other organisms by phage transduction, transposition or plasmid transfer and cis- or trans-complementation. For this, GMOs that are constructed with modern techniques of genetic engineering display a significant advantage over random mutagenesis derived live organisms. The selection of suitable GMO candidate strains can be made under in vitro conditions using basic knowledge on molecular mechanisms of pathogenicity of the corresponding bacterial species rather than by in vivo testing of large numbers of random mutants. This leads to a more targeted safety testing on volunteers and to a reduction in the use of animal experimentation.

Mechanisms contributing to the deep chlorophyll maximum in Lake Superior

The seasonal appearance of a deep chlorophyll maximum (DCM) in Lake Superior is a striking phenomenon that is widely observed; however its mechanisms of formation and maintenance are not well understood. As this phenomenon may be the reflection of an ecological driver, or a driver itself, a lack of understanding its driving forces limits the ability to accurately predict and manage changes in this ecosystem. Key mechanisms generally associated with DCM dynamics (i.e. ecological, physiological and physical phenomena) are examined individually and in concert to establish their role. First the prevailing paradigm, “the DCM is a great place to live”, is analyzed through an integration of the results of laboratory experiments and field measurements. The analysis indicates that growth at this depth is severely restricted and thus not able to explain the full magnitude of this phenomenon. Additional contributing mechanisms like photoadaptation, settling and grazing are reviewed with a one-dimensional mathematical model of chlorophyll and particulate organic carbon. Settling has the strongest impact on the formation and maintenance of the DCM, transporting biomass to the metalimnion and resulting in the accumulation of algae, i.e. a peak in the particulate organic carbon profile. Subsequently, shade adaptation becomes manifest as a chlorophyll maximum deeper in the water column where light conditions particularly favor the process. Shade adaptation mediates the magnitude, shape and vertical position of the chlorophyll peak. Growth at DCM depth shows only a marginal contribution, while grazing has an adverse effect on the extent of the DCM. The observed separation of the carbon biomass and chlorophyll maximum should caution scientists to equate the DCM with a large nutrient pool that is available to higher trophic levels. The ecological significance of the DCM should not be separated from the underlying carbon dynamics. When evaluated in its entirety, the DCM becomes the projected image of a structure that remains elusive to measure but represents the foundation of all higher trophic levels. These results also offer guidance in examine ecosystem perturbations such as climate change. For example, warming would be expected to prolong the period of thermal stratification, extending the late summer period of suboptimal (phosphorus-limited) growth and attendant transport of phytoplankton to the metalimnion. This reduction in epilimnetic algal production would decrease the supply of algae to the metalimnion, possibly reducing the supply of prey to the grazer community. This work demonstrates the value of modeling to challenge and advance our understanding of ecosystem dynamics, steps vital to reliable testing of management alternatives.

Opinion of headhunters about the ability of strabismic subjects to obtain employment

BACKGROUND: Through the opinion of Swiss headhunters, we wanted to determine the influence of strabismus on the ability to obtain employment. METHODS: Out of 31 randomly selected Swiss headhunters, 20 could be interviewed using a validated questionnaire. RESULTS: Forty-seven percent of the headhunters judged that strabismic subjects have more difficulties in obtaining a job. Gender has no influence on discrimination (p > 0.1). Asked about six facial disfigurements, strabismus was found to have the second largest negative impact on employment directly after acne. Strabismus was estimated to decrease the attractiveness of job applicants (p < 0.0001) and to have a negative impact on the overall judgment of a potential employer (p < 0.05). CONCLUSIONS: Visible strabismus influences negatively the ability to obtain a job. Because of its impact on the employability of a person, we believe that strabismus surgery in adults cannot be considered to be only a beautifying procedure.

Species-specific stomatal response of trees to drought - a link to vegetation dynamics?

Question: Is stomatal regulation specific for climate and tree species, and does it reveal species-specific responses to drought? Is there a link to vegetation dynamics? Location: Dry inner alpine valley, Switzerland Methods: Stomatal aperture (θE) of Pinus sylvestris, Quercus pubescens, Juniperus communis and Picea abies were continuously estimated by the ratio of measured branch sap flow rates to potential transpiration rates (adapted Penman-Monteith single leaf approach) at 10-min intervals over four seasons. Results: θE proved to be specific for climate and species and revealed distinctly different drought responses: Pinus stomata close disproportionately more than neighbouring species under dry conditions, but has a higher θE than the other species when weather was relatively wet and cool. Quercus keeps stomata more open under drought stress but has a lower θE under humid conditions. Juniperus was most drought-tolerant, whereas Picea stomata close almost completely during summer. Conclusions: The distinct microclimatic preferences of the four tree species in terms of θE strongly suggest that climate (change) is altering tree physiological performances and thus species-specific competitiveness. Picea and Pinus currently live at the physiological limit of their ability to withstand increasing temperature and drought intensities at the sites investigated, whereas Quercus and Juniperus perform distinctly better. This corresponds, at least partially, with regional vegetation dynamics: Pinus has strongly declined, whereas Quercus has significantly increased in abundance in the past 30 years. We conclude that θE provides an indication of a species' ability to cope with current and predicted climate.

Assessment of BoviPureTM for the In Vitro Production of Bovine Embryos

The objective of this study was to examine the potential utility of a commercially available sperm separation and purification product for the in vitro production of bovine embryos. Bovine oocytes were purchased from a commercial supplier, and matured oocytes were randomly allocated to one of two treatments. Oocytes were co-incubated with frozen-thawed semen washed twice with BoviPureTM (BoviPure group) or with modified Brackett-Oliphant medium (control group). After a 6-hour insemination period, oocytes were cultured in vitro for 8 days. Cleavage rate of embryos was determined 48 hours post-insemination, and blastocyst formation rate was assessed on day 8 of culture. The experiment was replicated three times, and data were analyzed using chi-square analysis. Washing of sperm in BoviPureTM had no effect (P>.05) on either cleavage rate (77.2%) or blastocyst development (21.6%) when compared with controls (71.9% and 17.1%, respectively). These results indicate that, under conditions of our study, the washing of sperm with BoviPureTM did not significantly enhance the ability to produce bovine embryos in vitro.

Grandparents in Kinship Care: Help or Hindrance to Family Preservation

The phenomenon of grandparents and other relatives raising children is a tradition rooted in the African American culture. However, a substantial increase in the number of relatives raising children has drawn attention to the child welfare system. Many of the biological parents are incarcerated for drugs or suffering from other social ills. Kinship care is an important component of family preservation and prevents court intervention based on child protection concerns and avoids formal placement of children in the child welfare system (Wilkerson, 1999). The child welfare system, however, is not conducive to this phenomenon. Placing children with grandparents and relatives allows them to live with people they know and trust; reduces the initial trauma of living with unknown persons; supports the transmission of identity, culture, and ethnicity; facilitates connections with brothers and sisters, and strengthens a family’s ability to provide the support they need.

Npro of classical swine fever virus contributes to pathogenicity in pigs by preventing type I interferon induction at local replication sites

Classical swine fever (CSF) caused by CSF virus (CSFV) is a highly contagious disease of pigs. The viral protein Npro of CSFV interferes with alpha- and beta-interferon (IFN-α/β) induction by promoting the degradation of interferon regulatory factor 3 (IRF3). During the establishment of the live attenuated CSF vaccine strain GPE-, Npro acquired a mutation that abolished its capacity to bind and degrade IRF3, rendering it unable to prevent IFN-α/β induction. In a previous study, we showed that the GPE- vaccine virus became pathogenic after forced serial passages in pigs, which was attributed to the amino acid substitutions T830A in the viral proteins E2 and V2475A and A2563V in NS4B. Interestingly, during the re-adaptation of the GPE- vaccine virus in pigs, the IRF3-degrading function of Npro was not recovered. Therefore, we examined whether restoring the ability of Npro to block IFN-α/β induction of both the avirulent and moderately virulent GPE--derived virus would enhance pathogenicity in pigs. Viruses carrying the N136D substitution in Npro regained the ability to degrade IRF3 and suppress IFN-α/β induction in vitro. In pigs, functional Npro significantly reduced the local IFN-α mRNA expression in lymphoid organs while it increased quantities of IFN-α/β in the circulation, and enhanced pathogenicity of the moderately virulent virus. In conclusion, the present study demonstrates that functional Npro influences the innate immune response at local sites of virus replication in pigs and contributes to pathogenicity of CSFV in synergy with viral replication.

Ontogenetic Effects of Hatching Plasticity in the Spotted Salamander (Ambystoma maculatum) due to Egg and Larval Predators

The ability to respond plastically to the environment has allowed amphibians to evolve a response to spatial and temporal variation in predation threat (Benard 2004). Embroys exposed to egg predation are expected to hatch out earlier than their conspecifics. Larval predation can induce a suite of phenotypic changes including growing a larger tail area. When presented with cues from both egg and larval predators, embryos are expected to respond to the egg predator by hatching out earlier because the egg predator presents an immediate threat. However, hatching early may be costly in the larval environment in terms of development, morphology, and/or behavior. We created a laboratory experiment in which we exposed clutches of spotted salamander (Ambystoma maculatum) eggs to both egg (caddisfly larvae) and larval (A. opacum) predators to test this hypothesis. We recorded hatching time and stage and took developmental and morphological data of the animals a week after hatching. Larvae were entered into lethal predation trials with a larval predatory sunfish (Lepomis sp.) in order to study behavior. We found that animals exposed to the egg predator cues hatched out earlier and at earlier developmental stages than conspecifics regardless of whether there was a larval predator present. Animals exposed to larval predator cues grew relatively larger tails and survived longer in the lethal predation trials. However the group exposed to both predators showed a cost of early hatching in terms of lower tail area and shorter survival time in predation trials. The morphological and developmental effects measured of hatching plasticity were transient as there were no developmental or morphological differences between the treatment groups at metamorphosis. Hatching plasticity may be transient but it is important to the development and survival of many amphibians.