Contrasting morphological and DNA barcode-suggested species boundaries among shallow-water amphipod fauna from the southern European Atlantic coast.

In this study we compared DNA barcode-suggested species boundaries with morphology-based species identifications in the amphipod fauna of the southern European Atlantic coast. DNA sequences of the cytochrome c oxidase subunit I barcode region (COI-5P) were generated for 43 morphospecies (178 specimens) collected along the Portuguese coast which, together with publicly available COI-5P sequences, produced a final dataset comprising 68 morphospecies and 295 sequences. Seventy-five BINs (Barcode Index Numbers) were assigned to these morphospecies, of which 48 were concordant (i.e., 1 BIN = 1 species), 8 were taxonomically discordant, and 19 were singletons. Twelve species had matching sequences (<2% distance) with conspecifics from distant locations (e.g., North Sea). Seven morphospecies were assigned to multiple, and highly divergent, BINs, including specimens of Corophium multisetosum (18% divergence) and Dexamine spiniventris (16% divergence), which originated from sampling locations on the west coast of Portugal (only about 36 and 250 km apart, respectively). We also found deep divergence (4%-22%) among specimens of seven species from Portugal compared to those from the North Sea and Italy. The detection of evolutionarily meaningful divergence among populations of several amphipod species from southern Europe reinforces the need for a comprehensive re-assessment of the diversity of this faunal group.

Broodstock diet effect on sea urchin Paracentrotus lividus (Lamarck, 1816) endotrophic larvae development: potential for their year-round use in environmental toxicology assessment.

The effect of captive broodstock diet on fertilization and endotrophic larvae development of the sea urchin Paracentrotus lividus was assessed. Maize grain and five inert pelleted diets were tested, during a three-month experimental period. Maize flour, wheat flour, soybean flour, maize/wheat flour (MWF) and maize/soybean flour mixes were used as vegetal sources for inert feed. Gonad index, percent egg fertilization and larvae malformation occurrence were compared with the results obtained from wild sea urchins (W). Whole egg total amino acid composition was concomitantly analyzed as a tool to explain eventual endotrophic larvae malformations caused by lack of specific nutrients. For all treatment groups (wild and captive), percent egg fertilization values above 96% were always observed, fulfilling the requisites (70-90%) necessary to conduct environmental monitoring bioassays, according to USEPA (2002). Similar values for normal percent larval development were only obtained from P. lividus broodstock subjected to an inert feeding diet based on a maize/wheat flour mix (85.0±1.45%), in comparison to wild P. lividus (82.5±1.75%). Likewise, no statistical differences on resultant whole egg total amino acid composition were observed between P. lividus fed MWF and wild treatments. Moreover, statistical differences between MWF and all the other captive feeding treatments were found for six out of the seventeen amino acids analyzed. This study demonstrates the possibility to obtain high values for P. lividus endotrophic larvae percent normal development based on broodstock held in captivity as long as an appropriate inert diet is provided.

DNA damage and metal accumulation in four tissues of feral Octopus vulgaris from two coastal areas in Portugal.

The alkaline comet assay has been employed for the first time to estimate the basal DNA damage in the digestive gland, gills, kidney and gonads of Octopus vulgaris. Octopuses were captured in two coastal areas adjacent to the cities of Matosinhos (N) and Olhão (S), Portugal. The area of Matosinhos is influenced by discharges of the Douro River, city of Porto, industries and intensive agriculture, while Olhão is an important fisheries port. Previous works point to contrasting metal availability in the two coastal areas. Among the analysed tissues digestive gland presented the highest levels of Zn, Cu, Cd and Pb. Tissues of specimens from Matosinhos exhibited high levels of Cd and from Olhão enhanced Pb concentrations. The DNA damages in digestive gland, gills and kidney were more accentuated in specimens from Matosinhos than from Olhão, suggesting a stronger effect of contaminants. Elevated strand breakages were registered in digestive gland, recognised for its ability to store and detoxify accumulated metals. The DNA damages in kidney, gills and gonads were lower, reflecting reduced metal accumulation or efficient detoxification. The broad variability of damages in the three tissues may also mirror tissue function, specific defences to genotoxicants and cell-cycle turnover.

Silica-based cationic bilayers as immunoadjuvants.

BACKGROUND: Silica particles cationized by dioctadecyldimethylammonium bromide (DODAB) bilayer were previously described. This work shows the efficiency of these particulates for antigen adsorption and presentation to the immune system and proves the concept that silica-based cationic bilayers exhibit better performance than alum regarding colloid stability and cellular immune responses for vaccine design. RESULTS: Firstly, the silica/DODAB assembly was characterized at 1 mM NaCl, pH 6.3 or 5 mM Tris.HCl, pH 7.4 and 0.1 mg/ml silica over a range of DODAB concentrations (0.001-1 mM) by means of dynamic light scattering for particle sizing and zeta-potential analysis. 0.05 mM DODAB is enough to produce cationic bilayer-covered particles with good colloid stability. Secondly, conditions for maximal adsorption of bovine serum albumin (BSA) or a recombinant, heat-shock protein from Mycobacterium leprae (18 kDa-hsp) onto DODAB-covered or onto bare silica were determined. At maximal antigen adsorption, cellular immune responses in vivo from delayed-type hypersensitivity reactions determined by foot-pad swelling tests (DTH) and cytokines analysis evidenced the superior performance of the silica/DODAB adjuvant as compared to alum or antigens alone whereas humoral response from IgG in serum was equal to the one elicited by alum as adjuvant. CONCLUSION: Cationized silica is a biocompatible, inexpensive, easily prepared and possibly general immunoadjuvant for antigen presentation which displays higher colloid stability than alum, better performance regarding cellular immune responses and employs very low, micromolar doses of cationic and toxic synthetic lipid.

Evaluation of a diphtheria and tetanus PLGA microencapsulated vaccine formulation without stabilizers.

Polymeric microspheres containing diphtheria and tetanus toxoids were prepared without protein stabilizers. A vaccine containing 2 Lf(tetanus) and 0.4 Lf(diphtheria) was injected either in BALB/c mice or in guinea-pigs. As control, a group received the alum-adsorbed unencapsulated toxoids. In mice, on day 44 one group and control received a booster and at day 111 the other group received the same booster dose. Before de booster, all groups had very low neutralizing antibodies, as determined by Toxin binding inhibition assay. One week after booster all groups had high antibody titers, especially those immunized with microencapsulated vaccine, which were at least 5 times higher than those immunized with alum vaccine for both antigens. Besides, guinea pigs receiving lower dose had antibodies titers as high as 60 UI/mL, and 30 times higher than those immunized with alum vaccine. Therefore by using an encapsulated vaccine without any kind of protein stabilizer we were able to induce in vivo protective responses irrespective of observed in vitro protein degradation by HPLC. Manipulating the vaccination schedule at the same time to the toxoids encapsulation does not only increase the antibody titers but also their specificity.

Ecological risk assessment of sediment management areas: application to Sado Estuary, Portugal.

The purpose of this work was to integrate different methodologies to assess the potential ecological risk of estuarine sedimentary management areas, using the Sado Estuary in Portugal as case study. To evaluate the environmental risk of sediment contamination, an integrative and innovative approach was used involving assessment of sediment chemistry, sediment toxicity, benthic community structure, human driving forces and pressures and management areas organic load levels. The basis for decision-making for overall assessment was a statistical multivariate analysis appended into a score matrix tables, using a best expert judgment. The integrated approach allowed to identify from the 19 management areas analyzed, three with no risk but other three with high risk to cause adverse effects in the biota, related with the contaminants analyzed. The methodologies used showed to be effective as a support for decision making leading to future estuarine management recommendations.

Nasopharyngeal carriage of Streptococcus pneumoniae among HIV-infected and -uninfected children <5 years of age before introduction of pneumococcal conjugate vaccine in Mozambique.

Nasopharyngeal carriage is a precursor for pneumococcal disease and can be useful for evaluating pneumococcal conjugate vaccine (PCV) impact. We studied pre-PCV pneumococcal carriage among HIV-infected and -uninfected children in Mozambique. Between October 2012 and March 2013, we enrolled HIV-infected children age <5 years presenting for routine care at seven HIV clinics in 3 sites, including Maputo (urban-south), Nampula (urban-north), and Manhiça (rural-south). We also enrolled a random sample of HIV-uninfected children <5 years old from a demographic surveillance site in Manhiça. A single nasopharyngeal swab was obtained and cultured following enrichment in Todd Hewitt broth with yeast extract and rabbit serum. Pneumococcal isolates were serotyped by Quellung reaction and multiplex polymerase chain reaction. Factors associated with pneumococcal carriage were examined using logistic regression. Overall pneumococcal carriage prevalence was 80.5% (585/727), with similar prevalences among HIV-infected (81.5%, 339/416) and HIV-uninfected (79.1%, 246/311) children, and across age strata. Among HIV-infected, after adjusting for recent antibiotic use and hospitalization, there was no significant association between study site and colonization: Maputo (74.8%, 92/123), Nampula (83.7%, 82/98), Manhiça (84.6%, 165/195). Among HIV-uninfected, report of having been born to an HIV-infected mother was not associated with colonization. Among 601 pneumococcal isolates from 585 children, serotypes 19F (13.5%), 23F (13.1%), 6A (9.2%), 6B (6.2%) and 19A (5.2%) were most common. The proportion of serotypes included in the 10- and 13-valent vaccines was 44.9% and 61.7%, respectively, with no significant differences by HIV status or age group. Overall 36.9% (n = 268) of children were colonized with a PCV10 serotype and 49.7% (n = 361) with a PCV13 serotype. Pneumococcal carriage was common, with little variation by geographic region, age, or HIV status. PCV10 was introduced in April 2013; ongoing carriage studies will examine the benefits of PCV10 among HIV-infected and-uninfected children.

Toxicity ranking of estuarine sediments on the basis of Sparus aurata biomarkers.

Sparus aurata biomarkers were used to rank sediments from the Sado River estuary (Portugal) according to their toxicity. Initially, the activities of liver ethoxyresorufin-O-deethylase, liver and gill glutathione S-transferases, muscle lactate dehydrogenase, and brain acetylcholinesterase were tested in a laboratory bioassay with the reference compound benzo[a]pyrene. Enzymatic activities were determined in different tissues of fish exposed for 48, 96, or 240 h to three concentrations of benzo[a]pyrene (25, 50, and 100 microg/L). Induction of liver ethoxyresorufin-O-deethylase was observed at all the exposure periods and concentrations, suggesting a continuous response of this system to toxicant exposure. Induction of liver glutathione S-transferases activity was only observed after 240 h of exposure, whereas gill glutathione S-transferases activity was significantly inhibited at all the exposure periods, suggesting a direct or indirect effect of the toxicant on these enzymes. Inhibition of lactate dehydrogenases activity was only observed after 96 h of exposure to 25 microg/L of benzo[a]pyrene. No significant effects were observed on acetylcholinesterase activity, suggesting that cholinergic function of S. aurata is not affected by benzo[a]pyrene. In a second phase, fish were exposed for 240 h to sediments collected at five sites of the Sado River estuary, and the same biomarkers were analyzed. For all the enzymes assayed, significant differences among sites were found. In this study, the battery of biomarkers used allowed to discrimination among sites with different types of contamination, levels of contamination, or both, after multivariate data analysis. Discrimination of sites was similar to the ranking provided by a more complex and parallel study (including chemical analysis of sediments, macrobenthic community analysis, amphipod mortality toxicity tests, and sea urchin abnormality embryo assays), suggesting its suitability to evaluate the toxicity of estuarine sediments.

DNA metabarcoding for high-throughput monitoring of estuarine macrobenthic communities.

Morphology-based profiling of benthic communities has been extensively applied to aquatic ecosystems' health assessment. However, it remains a low-throughput, and sometimes ambiguous, procedure. Despite DNA metabarcoding has been applied to marine benthos, a comprehensive approach providing species-level identifications for estuarine macrobenthos is still lacking. Here we report a combination of experimental and field studies to assess the aptitude of COI metabarcoding to provide robust species-level identifications for high-throughput monitoring of estuarine macrobenthos. To investigate the ability of metabarcoding to detect all species present in bulk DNA extracts, we contrived three phylogenetically diverse communities, and applied four different primer pairs to generate PCR products within the COI barcode region. Between 78-83% of the species in the contrived communities were recovered through HTS. Subsequently, we compared morphology and metabarcoding-based approaches to determine the species composition from four distinct estuarine sites. Our results indicate that species richness would be considerably underestimated if only morphological methods were used: globally 27 species identified through morphology versus 61 detected by metabarcoding. Although further refinement is required to improve efficiency and output of this approach, here we show the great aptitude of COI metabarcoding to provide high quality and auditable species identifications in estuarine macrobenthos monitoring.

Poly-lactide-co-glycolide microparticle sizes: a rational factorial design and surface response analysis.

Microsphere size is a primary determinant of solute release velocity. We present here a rational way for producing PLGA microspheres with different and controlled sizes. The following process variables were studied: Stirring velocity during the second emulsion step, dispersed and continuous phases volume ratio, and poly(vinyl alcohol) concentration in the continuous phase. A full factorial experimental design 2(3) with triplicate at the central point was used to determine the influence of variables on PLGA microsphere mean size. The stirring velocity and poly(vinyl alcohol) concentration were the main variables at 0.95 significance level. An influence of PVA and stirring velocity on microspheres size is observed, there is no correlation for DP/CP volume ratio on size of microspheres. By combining the two variables--the stirring velocity and poly(vinyl alcohol) concentration, the surface response was analyzed. The increase of poly(vinyl alcohol) concentration with concomitant increase on stirring velocity produced microspheres with the lower sized. In contrast the lower poly(vinyl alcohol) concentration and the lower stirring velocity used produced the higher microspheres sized. Uniformly spherical and smooth microspheres (4-15 microm of diameter) were obtained. No significant difference was observed on Ponca S loading within the experimental region. Our results open the possibility of formulating PLGA microspheres with custom sizes performing a minimum of experiments as required for specific applications.

Multi-level assessment of chronic toxicity of estuarine sediments with the amphipod Gammarus locusta: II. Organism and population-level endpoints.

This study aimed to test the performance of the amphipod Gammarus locusta (L.) in chronic sediment toxicity tests. It constitutes part of a multi-level assessment of chronic toxicity of estuarine sediments, integrating organism and population-level endpoints with biochemical markers responses. Here we account for organism and population-level effects, while biomarker responses were reported in a companion article. Five moderately contaminated sediments from Sado and Tagus estuaries were tested, comprising 3 muddy and 2 sandy sediments. These sediments either did not show acute toxicity or were diluted with control sediment as much as required to remove acute toxicity. Subsequent chronic tests consisted of 28-day exposures with survival, individual growth and reproductive traits as endpoints. Two of the muddy sediments induced higher growth rates in the amphipods, and improved reproductive traits. This was understood to be a consequence of the amount of organic matter in the sediment, which was nutritionally beneficial to the amphipods, while concurrently decreasing contaminant bioavailability. Biomarker responses did not reveal toxicant-induced stress in amphipods exposed to these sediments. One of the sandy sediments was acutely toxic at 50% dilution, but in contrast stimulated amphipod growth when diluted 75%. This was presumed to be an indication of a hormetic response. Finally the two remaining contaminated sediments showed pronounced chronic toxicity, affecting survival and reproduction. The sex ratio of survivors was highly biased towards females, and offspring production was severely impaired. The particulars of the responses of this amphipod were examined, as well as strengths versus limitations of the sediment test. This study illustrates the utility of this chronic test for toxicity assessment of contaminated estuarine sediments, with potential application all along Atlantic Europe.

Multi-level assessment of chronic toxicity of estuarine sediments with the amphipod Gammarus locusta: I. Biochemical endpoints.

We report on biomarker responses conducted as part of a multi-level assessment of the chronic toxicity of estuarine sediments to the amphipod Gammarus locusta. A companion article accounts for organism and population-level effects. Five moderately contaminated sediments from two Portuguese estuaries, Sado and Tagus, were assessed. Three of them were muddy and two were sandy sediments. The objective was to assess sediments that were not acutely toxic. Three of the sediments met this criterion, the other two were diluted (50% and 75%) with clean sediment until acute toxicity was absent. Following 28-d exposures, the amphipods were analysed for whole-body metal bioaccumulation, metallothionein induction (MT), DNA strand breakage (SB) and lipid peroxidation (LP). Two of the muddy sediments did not cause chronic toxicity. These findings were consistent with responses at organism and population levels that showed higher growth rates and improvement of reproductive traits for amphipods exposed to these two sediments. Two other sediments, one muddy and one sandy, exhibited pronounced chronic toxicity, affecting SB, MT induction (in muddy sediment), survival and reproduction. Potential toxicants involved in these effects were identified. The last sandy sediment exhibited some loss of DNA integrity, however growth was also enhanced. Present results, together with the organism/population-level data, and also benthic communities information, were analysed under a weight-of-evidence approach. By providing evidence of exposure (or lack of it) to contaminants in sediments, the biomarkers here applied assisted in distinguishing toxicants' impacts in test organisms from the confounding influence of other geochemical features of the sediments.

A rational design for the nanoencapsulation of poisonous animal venoms in liposomes prepared with natural phospholipids.

Liposomes have been used since the 1970's to encapsulate drugs envisaging enhancement in efficacy and therapeutic index, avoidance of side effects and increase in the encapsulated agent stability. The major problem when encapsulating snake venoms is the liposomal membrane instability caused by venom phospholipases. Here the results obtained encapsulating Crotalus durissimus terrificus and a pool of Bothropic venoms within liposomes (LC and LB, respectively) used to produce anti-venom sera are presented. The strategy was to modify the immunization protocol to enhance antibody production and to minimize toxic effects by encapsulating inactivated venoms within stabilized liposomes. Chemically modified venoms were solubilized in a buffer containing an inhibitor and a chelating agent. The structures of the venoms were analyzed by UV, CD spectroscopy and ELISA. In spite of the differences in the helical content between natural and modified venoms, they were recognized by horse anti-sera. To maintain long-term stability, mannitol was used as a cryoprotectant. The encapsulation efficiencies were 59 % (LB) and 99 % (LC), as followed by filtration on Sephacryl S1000. Light scattering measurements led us to conclude that both, LB (119 ±47 nm) and LC (147±56 nm) were stable for 22 days at 4 °C, even after lyophilization. Genetically selected mice and mixed breed horses were immunized with these formulations. The animals did not show clinical symptoms of venom toxicity. Both, LB and LC enhanced by at least 30 % the antibody titers 25 days after injection and total IgG titers remained high 91 days after immunization. The liposomal formulation clearly exhibited adjuvant properties.

Risco e Sinthome: A Psicanálise no Sistema Socioeducativo / Risk and Sinthome: Psychoanalysis in the Socio-educational System

Pretendemos discutir a aplicação da teoria psicanalítica às políticas públicas voltadas à socioeducação de adolescentes autores de ato infracional. Partimos de revisão sobre adolescência e clínica psicanalítica, especialmente com adolescentes infratores. Com discussão de caso de uma adolescente em cumprimento de medida socioeducativa de liberdade assistida, levantamos duas considerações sobre a psicanálise nesse dispositivo: a singularidade do caso face ao universal da regulação jurídica e os limites e as potencialidades do atendimento institucional. Concluímos pela importância da palavra e pelo recolhimento de seu excedente no trabalho subjetivo do adolescente, verificando a necessidade de ações que encaminhem os efeitos das intervenções. Podemos dizer que o efeito político-social esperado pela aplicação de uma medida socioeducativa não caminha sem a consideração do mais singular e íntimo de cada adolescente, do qual sua posição subjetiva faz testemunho, delimitando seu modo de fazer exceção à regra e habitar o mundo.

We intend to discuss the application of psychoanalytic theory to public policies towards delinquent adolescents. We start from a review of adolescence and psychoanalytic practice, especially with young offenders. With a case of a teenager in under socio-educational measure in assisted liberty, two considerations raised about psychoanalysis in this service: the uniqueness of the case against the universal legal regulation, as well as limitations and potential of institutional care. We conclude by the importance of the word and the gathering surplus in the subjective labor of the adolescent and finally, we see the need for actions that forward the effects of interventions. We state that the expected social-political effect of applying an educational measure needs to consider the uniqueness of each teenager, of which his or her subjective position is testimony, delimiting a way of being an exception to the rule and inhabiting the world.

[Laparoscopic treatment of liver cysts]. / Tratamento videolaparoscópico do cisto hepático.

OBJECTIVE: The authors present their experience with the laparoscopic management of non-parasitic liver cysts. METHODS: During the period from May 2003 to August 2006, thirteen patients with non-parasitic liver cysts underwent laparoscopic fenestration. Eleven patients were female (84.6%) and two (15.4%) were male. The average age at diagnosis was 48.3 years (range 35-72). The majority of patients presented with pain, discomfort, dyspepsia, or early satiety at diagnosis. Computed tomography scans were performed in all cases. The cysts were solitary in eleven patients (84.6%) and polycystic liver disease in two patients (15.4%). The average size of the solitary cysts was 11.3 cm (range 9.5-17 cm), and the polycystic liver disease was 10.6 cm (range 9.2-12.1 cm). Operation was indicated due to symptoms in all patients. The technique performed was laparoscopic cyst fenestration. RESULTS: Laparoscopic fenestration was completed in all patients. Mean operative time was 85 minutes (range 53-110). Intraoperative complications were not detected and there was no death. Postoperative complications were observed in two patients (15.4%). Bile leakage was present in one patient, and ascites in another patient. Both cases were treated conservatively. The average hospital stay was 3.5 days (range 2-9 days). Mean follow-up was 36 months. There was one asymptomatic recurrence (7.6%) in a patient with polycystic liver disease after two years of follow-up. CONCLUSION: Laparoscopic fenestration is the preferred method of treatment for non-parasitic liver cyst. This operation causes low morbidity and the recurrence is uncommon. Adequate selection of patients and the laparoscopic technique are important for the treatment.

Novel immunoadjuvants based on cationic lipid: Preparation, characterization and activity in vivo.

The interactions between three different protein antigens and dioctadecyldimethylammonium bromide (DODAB) dispersed in aqueous solutions from probe sonication or adsorbed as one bilayer onto particles was comparatively investigated. The three model proteins were bovine serum albumin (BSA), purified 18 kDa/14 kDa antigens from Taenia crassiceps (18/14-Tcra) and a recombinant, heat-shock protein hsp-18 kDa from Mycobacterium leprae. Protein-DODAB complexes in water solution were characterized by dynamic light scattering for sizing and zeta-potential analysis. Cationic complexes (80-100 nm of mean hydrodynamic diameter) displayed sizes similar to those of DODAB bilayer fragments (BF) in aqueous solution and good colloid stability over a range of DODAB and protein concentrations. The amount of cationic lipid required for attaining zero of zeta-potential at a given protein amount depended on protein nature being smaller for 18 kDa/14 kDa antigens than for BSA. Mean diameters for DODAB/protein complexes increased, whereas zeta-potentials decreased with NaCl or protein concentration. In mice, weak IgG production but significant cellular immune responses were induced by the complexes in comparison to antigens alone or carried by aluminum hydroxide as shown from IgG in serum determined by ELISA, delayed type hypersensitivity reaction from footpad swelling tests and cytokines analysis. The novel cationic adjuvant/protein complexes revealed good colloid stability and potential for vaccine design at a reduced DODAB concentration.

LUVs recovered with Chitosan: a new preparation for vaccine delivery.

Chitosan, alpha-(1-4)-amino-2-deoxy-beta-D-glucan, is a deacetylated form of chitin, an abundant natural polysaccharide present in crustacean shells. Its unique characteristics such as positive charge, biodegradability, biocompatibility, nontoxicity, and rigid linear molecular structure make this macromolecule ideal as drug carrier. The association between chitosan and liposomes was carefully described, where REVs (reverse phase evaporation vesicles) were sandwiched by chitosan. The usage of these particles in vaccine formulation is here proposed for the first time in the literature. The Chitosan-REVs now stabilized by polyvinilic alcohol were the vehicle for Diphtheria toxoid (Dtxd). Round chitosan-sandwiched REVs (REVs-Chi) particles of 373 +/- 17 nm containing 65% Dtxd were obtained. After 200 min of incubation in a simulated gastric fluid, 70% of the Dtxd was liberated from REVs-Chi in comparison to 100% of Dtxd liberated from pure REVs. In PBS, the Dtxd liberation from REVS-Chi was about 60%. Mice were immunized with Dtxd encapsulated within REVs-Chi and with other REVs/Dtxd formulations adsorbed onto Freund adjuvant or alumen [AIF and Al(OH)(3)]. The response patterns and the immune maturity were measured by IgG(1) and IgG(2a) titrations. REVs-Chi containing Dtxd elicited both antibodies production giving the animals higher immune response and selectivity. It was interesting that the memory of those mice immunized with REVs-Chi containing Dtxd enhanced, after booster, antibody production by 47% in contrast with 17 and 7% in mice immunized with the antigen vehiculated in REVs-AIF or REVs-Al(OH)(3), respectively.

Heat shock protein micro-encapsulation as a double tool for the improvement of new generation vaccines.

The modern vaccinology encompasses the recombinant DNA technology, protein and carbohydrate chemistry to obtain safe molecularly defined vaccines. Nevertheless most of the vaccines are poorly immunogenic because a large number of antigens are membrane proteins and consequently they are not present in their active conformation in the vaccine. Others are not as potent because they contain only B epitopes and therefore, cannot stimulate cellular memory. We have been studying the characteristics of the recombinant heat shock protein 18kDa-hsp from Mycobacterium leprae as an alternative carrier protein with a T epitope source to enhance the activity of these second generation vaccines. Here we proved that the 18kDa-hsp acted as carrier, without masking the activity of the carried antigen, with similar immune stimulatory effect when compared with ODN1668. Supramolecular aggregates of 18kDa-hsp and Mice serum albumin (MSA) were obtained using glutaraldehyde as cross linker. The Neisseria meningitides serogroup C polysaccharide (PSC, a B epitope) and the carrier protein 18kDa-hsp were co-encapsulated within Soybean phosphatidylcholine liposomes (SPC: Cho : alpha-Toc, 22 : 5 : 0.18 molar ratio, respectively). These liposomes were prepared in MPB buffer (20 mM phosphate, 295 mM mannitol pH 7.2) in the presence or absence of the ODN1668, TCCATGACGTTCCTGATGCT. When mice were injected with 18kDa-hsp-MSA no antibody against the MSA was observed. This means that the 18kDa-hsp acted as carrier, without masking the carried protein immune activity. Stable liposomes of 150 nm were obtained using mannitol as a cryoprotector. Genetically selected mice when injected with liposomes containing PSC and 18kDa-hsp displayed an antibody titer of 12. In contrast, in those mice injected with free PSC there was no response. The 18kDa-hsp adjuvant effect on the PSC liposomal formulation was comparable to that observed when ODN1668 was co-encapsulated with PSC. Confirming our expectations we observed that the formulation containing 18kDa-hsp conferred a memory response to the carried antigen--the Neisseria meningitidis serogroup C polysaccharide.

Tratamento videolaparoscópico do cisto hepático / Laparoscopic treatment of liver cysts

OBJETIVO: Os autores apresentam a experiência no tratamento laparoscópico do cisto hepático não parasitário. MÉTODOS: No período de maio de 2003 a agosto de 2006, treze pacientes com cisto hepático não parasitário foram submetidos a tratamento com fenestração laparoscópica. Havia dois pacientes do sexo masculino (15,4 por cento) e onze do sexo feminino (84,6 por cento), com idade média no momento do diagnóstico de 48,3 anos (variação de 35 a 72 anos). A maioria dos pacientes apresentava dor e desconforto abdominal, dispepsia e saciedade precoce ao diagnóstico. A tomografia computdorizada do abdome foi realizada em todos os pacientes e o cisto solitário diagnosticado em onze pacientes (84,6 por cento) e doença policística em dois pacientes (15,4 por cento). O tamanho médio do cisto solitário era 11,3 cm (variação de 9,5 a 17 cm) e doença policística era 10,6 cm (um paciente com o maior cisto de 9,2 cm e o outro com 12,1 cm). A cirurgia foi indicada devido a todos os pacientes se apresentarem sintomáticos e a técnica utilizada foi a fenestração laparoscópica. RESULTADOS: O procedimento de fenestração laparoscópica foi realizado com êxito em todos os pacientes. O tempo operatório médio foi de 85 min (variação de 53 a 110 minutos). Não ocorreu óbito ou complicações intra-operatórias. Complicações pós-operatórias foram observadas em dois pacientes (15,4 por cento). Fístula biliar em um paciente e ascite em outro paciente, sendo tratados de forma conservadora com sucesso. O tempo médio de internação hospitalar foi de 3,5 dias (variação de 2 a 9 dias) e o seguimento médio foi de 36 meses. Foi observada uma recorrência assintomática (7,6 por cento) em paciente com doença policística após dois anos de seguimento. CONCLUSÃO: A fenestração laparoscópica é o método preferido para o tratamento do cisto hepático não parasitário. Este procedimento apresenta baixa morbidade associada a recorrência incomum. A adequada seleção dos pacientes e a técnica ...

OBJECTIVE: The authors present their experience with the laparoscopic management of non-parasitic liver cysts. METHODS: During the period from May 2003 to August 2006, thirteen patients with non-parasitic liver cysts underwent laparoscopic fenestration. Eleven patients were female (84.6 percent) and two (15.4 percent) were male. The average age at diagnosis was 48.3 years (range 35-72). The majority of patients presented with pain, discomfort, dyspepsia, or early satiety at diagnosis. Computed tomography scans were performed in all cases. The cysts were solitary in eleven patients (84.6 percent) and polycystic liver disease in two patients (15.4 percent). The average size of the solitary cysts was 11.3 cm (range 9.5-17 cm), and the polycystic liver disease was 10.6 cm (range 9.2-12.1 cm). Operation was indicated due to symptoms in all patients. The technique performed was laparoscopic cyst fenestration. RESULTS: Laparoscopic fenestration was completed in all patients. Mean operative time was 85 minutes (range 53-110). Intraoperative complications were not detected and there was no death. Postoperative complications were observed in two patients (15.4 percent). Bile leakage was present in one patient, and ascites in another patient. Both cases were treated conservatively. The average hospital stay was 3.5 days (range 2-9 days). Mean follow-up was 36 months. There was one asymptomatic recurrence (7.6 percent) in a patient with polycystic liver disease after two years of follow-up. CONCLUSION: Laparoscopic fenestration is the preferred method of treatment for non-parasitic liver cyst. This operation causes low morbidity and the recurrence is uncommon. Adequate selection of patients and the laparoscopic technique are important for the treatment.