A sensitive flow cytometric methodology for studying the binding of L. chagasi to canine peritoneal macrophages

BACKGROUND: The Leishmania promastigote-macrophage interaction occurs through the association of multiple receptors on the biological membrane surfaces. The success of the parasite infection is dramatically dependent on this early interaction in the vertebrate host, which permits or not the development of the disease. In this study we propose a novel methodology using flow cytometry to study this interaction, and compare it with a previously described "in vitro" binding assay. METHODS: To study parasite-macrophage interaction, peritoneal macrophages were obtained from 4 dogs and adjusted to 3 × 10(6 )cells/mL. Leishmania (Leishmania) chagasi parasites (stationary-phase) were adjusted to 5 × 10(7 )cells/mL. The interaction between CFSE-stained Leishmania chagasi and canine peritoneal macrophages was performed in polypropylene tubes to avoid macrophage adhesion. We carried out assays in the presence or absence of normal serum or in the presence of a final concentration of 5% of C5 deficient (serum from AKR/J mice) mouse serum. Then, the number of infected macrophages was counted in an optical microscope, as well as by flow citometry. Macrophages obtained were stained with anti-CR3 (CD11b/CD18) antibodies and analyzed by flow citometry. RESULTS: Our results have shown that the interaction between Leishmania and macrophages can be measured by flow cytometry using the fluorescent dye CFSE to identify the Leishmania, and measuring simultaneously the expression of an important integrin involved in this interaction: the CD11b/CD18 (CR3 or Mac-1) β2 integrin. CONCLUSION: Flow cytometry offers rapid, reliable and sensitive measurements of single cell interactions with Leishmania in unstained or phenotypically defined cell populations following staining with one or more fluorochromes

Fetal deaths in Brazil: a systematic review

OBJECTIVE To review the frequency of and factors associated with fetal death in the Brazilian scientific literature. METHODS A systematic review of Brazilian studies on fetal deaths published between 2003 and 2013 was conducted. In total, 27 studies were analyzed; of these, 4 studies addressed the quality of data, 12 were descriptive studies, and 11 studies evaluated the factors associated with fetal death. The databases searched were PubMed and Lilacs, and data extraction and synthesis were independently performed by two or more examiners. RESULTS The level of completeness of fetal death certificates was deficient, both in the completion of variables, particularly sociodemographic variables, and in defining the underlying causes of death. Fetal deaths have decreased in Brazil; however, inequalities persist. Analysis of the causes of death indicated maternal morbidities that could be prevented and treated. The main factors associated with fetal deaths were absent or inadequate prenatal care, low education level, maternal morbidity, and adverse reproductive history. CONCLUSIONS Prenatal care should prioritize women that are most vulnerable (considering their social environment or their reproductive history and morbidities) with the aim of decreasing the fetal mortality rate in Brazil. Adequate completion of death certificates and investment in the committees that investigate fetal and infant deaths are necessary

Single-dose administration and the influence of the timing of the booster dose on immunogenicity and efficacy of ChAdOx1 nCoV-19 (AZD1222) vaccine: a pooled analysis of four randomised trials.

BACKGROUND: The ChAdOx1 nCoV-19 (AZD1222) vaccine has been approved for emergency use by the UK regulatory authority, Medicines and Healthcare products Regulatory Agency, with a regimen of two standard doses given with an interval of 4-12 weeks. The planned roll-out in the UK will involve vaccinating people in high-risk categories with their first dose immediately, and delivering the second dose 12 weeks later. Here, we provide both a further prespecified pooled analysis of trials of ChAdOx1 nCoV-19 and exploratory analyses of the impact on immunogenicity and efficacy of extending the interval between priming and booster doses. In addition, we show the immunogenicity and protection afforded by the first dose, before a booster dose has been offered. METHODS: We present data from three single-blind randomised controlled trials-one phase 1/2 study in the UK (COV001), one phase 2/3 study in the UK (COV002), and a phase 3 study in Brazil (COV003)-and one double-blind phase 1/2 study in South Africa (COV005). As previously described, individuals 18 years and older were randomly assigned 1:1 to receive two standard doses of ChAdOx1 nCoV-19 (5 × 1010 viral particles) or a control vaccine or saline placebo. In the UK trial, a subset of participants received a lower dose (2·2 × 1010 viral particles) of the ChAdOx1 nCoV-19 for the first dose. The primary outcome was virologically confirmed symptomatic COVID-19 disease, defined as a nucleic acid amplification test (NAAT)-positive swab combined with at least one qualifying symptom (fever ≥37·8°C, cough, shortness of breath, or anosmia or ageusia) more than 14 days after the second dose. Secondary efficacy analyses included cases occuring at least 22 days after the first dose. Antibody responses measured by immunoassay and by pseudovirus neutralisation were exploratory outcomes. All cases of COVID-19 with a NAAT-positive swab were adjudicated for inclusion in the analysis by a masked independent endpoint review committee. The primary analysis included all participants who were SARS-CoV-2 N protein seronegative at baseline, had had at least 14 days of follow-up after the second dose, and had no evidence of previous SARS-CoV-2 infection from NAAT swabs. Safety was assessed in all participants who received at least one dose. The four trials are registered at ISRCTN89951424 (COV003) and ClinicalTrials.gov, NCT04324606 (COV001), NCT04400838 (COV002), and NCT04444674 (COV005). FINDINGS: Between April 23 and Dec 6, 2020, 24 422 participants were recruited and vaccinated across the four studies, of whom 17 178 were included in the primary analysis (8597 receiving ChAdOx1 nCoV-19 and 8581 receiving control vaccine). The data cutoff for these analyses was Dec 7, 2020. 332 NAAT-positive infections met the primary endpoint of symptomatic infection more than 14 days after the second dose. Overall vaccine efficacy more than 14 days after the second dose was 66·7% (95% CI 57·4-74·0), with 84 (1·0%) cases in the 8597 participants in the ChAdOx1 nCoV-19 group and 248 (2·9%) in the 8581 participants in the control group. There were no hospital admissions for COVID-19 in the ChAdOx1 nCoV-19 group after the initial 21-day exclusion period, and 15 in the control group. 108 (0·9%) of 12 282 participants in the ChAdOx1 nCoV-19 group and 127 (1·1%) of 11 962 participants in the control group had serious adverse events. There were seven deaths considered unrelated to vaccination (two in the ChAdOx1 nCov-19 group and five in the control group), including one COVID-19-related death in one participant in the control group. Exploratory analyses showed that vaccine efficacy after a single standard dose of vaccine from day 22 to day 90 after vaccination was 76·0% (59·3-85·9). Our modelling analysis indicated that protection did not wane during this initial 3-month period. Similarly, antibody levels were maintained during this period with minimal waning by day 90 (geometric mean ratio [GMR] 0·66 [95% CI 0·59-0·74]). In the participants who received two standard doses, after the second dose, efficacy was higher in those with a longer prime-boost interval (vaccine efficacy 81·3% [95% CI 60·3-91·2] at ≥12 weeks) than in those with a short interval (vaccine efficacy 55·1% [33·0-69·9] at <6 weeks). These observations are supported by immunogenicity data that showed binding antibody responses more than two-fold higher after an interval of 12 or more weeks compared with an interval of less than 6 weeks in those who were aged 18-55 years (GMR 2·32 [2·01-2·68]). INTERPRETATION: The results of this primary analysis of two doses of ChAdOx1 nCoV-19 were consistent with those seen in the interim analysis of the trials and confirm that the vaccine is efficacious, with results varying by dose interval in exploratory analyses. A 3-month dose interval might have advantages over a programme with a short dose interval for roll-out of a pandemic vaccine to protect the largest number of individuals in the population as early as possible when supplies are scarce, while also improving protection after receiving a second dose. FUNDING: UK Research and Innovation, National Institutes of Health Research (NIHR), The Coalition for Epidemic Preparedness Innovations, the Bill & Melinda Gates Foundation, the Lemann Foundation, Rede D'Or, the Brava and Telles Foundation, NIHR Oxford Biomedical Research Centre, Thames Valley and South Midland's NIHR Clinical Research Network, and AstraZeneca

A percepção de profissionais da saúde de um hospital sobre a violência contra as mulheres La percepción de profesionales de salud de un hospital sobre la violencia contra la mujer The perceptions of the health professionals at a hospital about the violence against women

Estudo qualitativo que objetivou conhecer percepções de profissionais de saúde acerca da assistência às mulheres em situação de violência. O cenário foi um hospital de ensino no sul do Brasil. Os sujeitos da pesquisa foram 12 profissionais de saúde do Centro Obstétrico e Pronto Socorro. A coleta dos dados realizou-se em abril/maio de 2008, a partir de vinheta e entrevista semi-estruturada. As informações foram organizadas e estruturadas por meio da análise temática. Dos resultados emergiram duas categorias: o diálogo como instrumento facilitador para a identificação da violência e seus encaminhamentos, e a complexidade do fenômeno e suas implicações na assistência. Aponta-se a necessidade de criação de espaços de responsabilização dos órgãos formadores e serviços de saúde na formação de equipe multiprofissional que discuta e implemente a política de atendimento às mulheres em situação de violência.<br>Estudio cualitativo que tuvo por objetivo conocer las percepciones de profesionales de salud acerca de la asistencia a las mujeres en situaciones de violencia. El escenario fue un hospital de enseñanza en el sur de Brasil. Los sujetos de la investigación fueron 12 profesionales de salud del Centro Obstétrico y Hospital de urgencias. La recolección de los datos se realizó en abril/mayo de 2008, a partir de viñeta y entrevista semiestructurada. Las informaciones fueron organizadas y estructuradas por medio del análisis temático. Los resultados emergieron dos categorías: el diálogo como instrumento facilitador para la identificación de la violencia y sus encaminamientos las mujeres y la complexidad del fenómeno y sus implicaciones en la asistencia. Se evidencia la necesidad de creación de espacios de responsabilización de los órganos formadores y servicios de salud en la formación del equipo multiprofesional que discuta e implemente la política de atención a las mujeres en situación de violencia.<br>This qualitative study aimed to know the perceptions of health professionals about the assistance to women living in a violent situation. The scenery was a teaching hospital of southern Brazil. The participants were 12 health professionals from the obstetric and emergency units. The data were collected by semistructured interviews started by a short story in April to May of 2008. The information was organized and categorized and subject to thematic analysis. The results showed two categories: the dialogue as a facilitator instrument to identify the violence and their referrals and the complexity of the phenomena and their implications to the assistance. We pointed out the necessity of to create spaces of the responsibility of agencies devoted to training human resources in health and health services. It is in order to form multiprofessional team that discuss and implements the policy care to women living in a violent situation