Comparative evaluation of PCR in Ziehl-Neelsen stained smears and PCR in tissues for diagnosis of Mycobacterium avium subsp. paratuberculosis.

Thirty six tissues from sheep, previously diagnosed with paratuberculosis, were tested by PCR in positive Ziehl-Neelsen staining smears of tissues, and PCR in tissues targeting IS900 specific for Mycobacterium avium subsp. paratuberculosis. DNA amplification was achieved in 33.3% Ziehl-Neelsen smears, and in 61.1% tissue samples. Combination of both techniques found 66.7% samples as positive. Combination of techniques would, therefore, increase the sensitivity of diagnosis.

Bacterial diversity associated with the Brazilian endemic reef coral Mussismilia braziliensis.

AIMS: We performed the first characterization of the microbiota associated with the reef coral Mussismilia braziliensis by means of a culture-independent approach. METHODS AND RESULTS: The main groups were Proteobacteria, Cyanobacteria and unclassified bacteria according to the 16S rDNA libraries. Most of the sequences of the mucus of healthy and diseased M. braziliensis did not find close matches in GenBank (i.e. >97% 16S rDNA similarity). Most of the sequences of seawater and mucus of healthy coral fell into tight clusters (17 and 15 clusters respectively). In contrast, most of the sequences of mucus of diseased coral did not form clusters. The rarefaction curves indicate saturation in the recovery of higher taxa (approximately 40 phyla). However, the number of species in the coral mucus (n = 130-170) and seawater (n = 170) did not reach a plateau. CONCLUSIONS: The coral microbiota encompasses several potentially novel species and higher taxa. The microbiota of M. braziliensis appears to be species-specific. Diseased coral may have provided a suitable place for colonization by opportunistic bacteria, resulting in a greater bacterial diversity. SIGNIFICANCE AND IMPACT OF THE STUDY: The first study on the diversity of the microbiota of the endemic and endangered of extinction coral M. braziliensis.

Serological prevalence of toxoplasmosis and neosporosis in dogs diagnosed with suspected meningoencephalitis in the UK.

OBJECTIVES: To assess the prevalence of antibodies to Toxoplasma gondii and Neospora caninum in a population of dogs with a diagnosis of suspected inflammatory meningoencephalitis. MATERIALS AND METHODS: Medical records of three referral centres were reviewed from 2008 to 2016 to identify a cohort of dogs diagnosed and treated for suspected inflammatory meningoencephalitis after testing for evidence of exposure to these pathogens. RESULTS: In our sample of 400 dogs the prevalence for exposure (IgG>1:50) to Toxoplasma gondii was 8/201 (3∙98%). Active infection (IgG titre >1:400 or/and an IgM titre >1:64 and/or positive PCR in CSF) was suspected in 1/400 (0∙25%). The prevalence for exposure [Indirect fluorescent antibody (IFA) titre >1:50] and active infection (IFA titres ≥⃒1:400 and/or positive PCR in CSF) with Neospora caninum were 14/201 (6∙96%) and 9/400 (2∙25%), respectively. CLINICAL SIGNIFICANCE: In view of the low prevalence of protozoan infections, the risk associated with starting immunosuppressive medication in dogs with evidence of inflammatory meningitis or encephalitis in the UK appears low.

A case-control study of risk factors for brucellosis seropositivity in Portuguese small ruminants herds.

A case-control study involving 255 small ruminants herds randomly selected was carried out in Portugal between January and December 2004, to identify risk factors associated with brucellosis seropositivity. To achieve this objective, two groups of herds selected according their prevalence status were compared: "cases" (farms with seroprevalence higher than 5%, n=123) and "controls" (farms seronegatives, n=132). A carefully structured questionnaire was used to collect data from each herd. A statistical analysis to compare "case" versus "control" herds was performed with the variables obtained from the questionnaire and the seroprevalence results. The effects on seroprevalence of several variables such as: individual characteristics; farm management practices; farm characteristics; animal health; knowledge and characteristics of farmers were evaluated. Data were analysed using logistic regression. Univariable analysis was used to screen the variables used in the logistic regression model. Nine variables were associated with brucellosis seropositivity in univariable analysis p<0.10. These variables were retained for multivariable logistic regression model. Regression model identified five variables as risk factors for seropositivity. The odds of brucellosis were increased: herds with more than 116 animals (OR=2.99); in herds with no cleaned-watering places (OR=3.05); in herds with insufficient manure removal and insufficient cleaning of premises (OR=2.87); in introduction of animals from non-free brucellosis herds or from herds of unknown status (OR=12.11). In the other hand, farmers' age (the eldest) was related to decreased odds (OR=0.4). Potential risk factors identified in this study were consistent factors associated with brucellosis seropositivity and support current recommendations for the control of brucellosis. Considering the paucity of epidemiological reports on brucellosis in the Northeast of Portugal and the absence of any data concerning factors related to either the prevention or the spread of the disease, our results could make a useful contribution towards the prevention of small ruminants brucellosis in the area.

Liver mitochondrial dysfunction and oxidative stress in the pathogenesis of experimental nonalcoholic fatty liver disease.

Oxidative stress and hepatic mitochondria play a role in the pathogenesis of nonalcoholic fatty liver disease. The aim of the present study was to evaluate the role of hepatic mitochondrial dysfunction and oxidative stress in the pathogenesis of the disease. Fatty liver was induced in Wistar rats with a choline-deficient diet (CD; N = 7) or a high-fat diet enriched with PUFAs-omega-3 (H; N = 7) for 4 weeks. The control group (N = 7) was fed a standard diet. Liver mitochondrial oxidation and phosphorylation were measured polarographically and oxidative stress was estimated on the basis of malondialdehyde and glutathione concentrations. Moderate macrovacuolar liver steatosis was observed in the CD group and mild liver steatosis was observed in the periportal area in the H group. There was an increase in the oxygen consumption rate by liver mitochondria in respiratory state 4 (S4) and a decrease in respiratory control rate (RCR) in the CD group (S4: 32.70 +/- 3.35; RCR: 2.55 +/- 0.15 ng atoms of O2 min-1 mg protein-1) when compared to the H and control groups (S4: 23.09 +/- 1.53, 17.04 +/- 2.03, RCR: 3.15 +/- 0.15, 3.68 +/- 0.15 ng atoms of O2 min-1 mg protein-1, respectively), P < 0.05. Hepatic lipoperoxide concentrations were significantly increased and the concentration of reduced glutathione was significantly reduced in the CD group. A choline-deficient diet causes moderate steatosis with disruption of liver mitochondrial function and increased oxidative stress. These data suggest that lipid peroxidation products can impair the flow of electrons along the respiratory chain, causing overreduction of respiratory chain components and enhanced mitochondrial reactive oxygen species. These findings are important in the pathogenesis of nonalcoholic fatty liver disease.

Sorafenib prevents liver fibrosis in a non-alcoholic steatohepatitis (NASH) rodent model.

Liver fibrosis occurring as an outcome of non-alcoholic steatohepatitis (NASH) can precede the development of cirrhosis. We investigated the effects of sorafenib in preventing liver fibrosis in a rodent model of NASH. Adult Sprague-Dawley rats were fed a choline-deficient high-fat diet and exposed to diethylnitrosamine for 6 weeks. The NASH group (n=10) received vehicle and the sorafenib group (n=10) received 2.5 mg·kg(-1)·day(-1) by gavage. A control group (n=4) received only standard diet and vehicle. Following treatment, animals were sacrificed and liver tissue was collected for histologic examination, mRNA isolation, and analysis of mitochondrial function. Genes related to fibrosis (MMP9, TIMP1, TIMP2), oxidative stress (HSP60, HSP90, GST), and mitochondrial biogenesis (PGC1α) were evaluated by real-time quantitative polymerase chain reaction (RT-qPCR). Liver mitochondrial oxidation activity was measured by a polarographic method, and cytokines by enzyme-linked immunosorbent assay (ELISA). Sorafenib treatment restored mitochondrial function and reduced collagen deposition by nearly 63% compared to the NASH group. Sorafenib upregulated PGC1α and MMP9 and reduced TIMP1 and TIMP2 mRNA and IL-6 and IL-10 protein expression. There were no differences in HSP60, HSP90 and GST expression. Sorafenib modulated PGC1α expression, improved mitochondrial respiration and prevented collagen deposition. It may, therefore, be useful in the treatment of liver fibrosis in NASH.

Role of tachykinin NK2 receptors in normal and altered rectal sensitivity in rats.

Irritable bowel syndrome is characterized by visceral hyperalgesia commonly associated with stress and inflammatory processes. We investigated the role of tachykinin NK2 receptors in the ability of trinitrobenzenesulphonic acid (TNBS) and stress to enhance the sensitivity of the rat rectum to distension using a selective tachykinin NK2 receptor antagonist (MEN 11420). Rats were fitted with electrodes implanted in the striated muscles of the abdomen. Rectal distension (RD) was performed with a balloon inflated by steps of 0.4 ml from 0 to 1.6 ml. Five groups were submitted to RD performed 3 days before and after intrarectal instillation of TNBS. Fifteen minutes before RD, rats were treated with saline or MEN 11420 (5 - 100 microg kg(-1) i.v.). Two other groups, submitted to 2 h restraint or sham stress sessions were randomly treated i.v. with saline or MEN 11420 (10 - 200 microg kg(-1)) prior to RD applied 20 min later. The basal response to RD was characterized by a significant increase in the number of abdominal contractions. This response occurred with a threshold volume of 0.8 ml and was dose-dependently reduced by MEN 11420 (5 - 100 microg kg(-1) i.v.). Rectal inflammation lowered the volume of distension producing abdominal contractions to 0.4 ml (allodynia). This effect was either reduced or suppressed by MEN 11420. A similar allodynia was observed after a stress session and this effect was reduced (49%) or suppressed by MEN 11420 at 200 and 100 microg kg(-1), respectively. Tachykinin NK2 receptors are involved in rectal hypersensitivity associated with inflammation and stress. British Journal of Pharmacology (2000) 129, 193 - 199

Detection of Vibrio cholerae and V. mimicus heat-stable toxin gene sequence by PCR.

Previously the heat-stable enterotoxin in Vibrio cholerae and V. mimicus has been detected by suckling mouse assay, a non-specific approach, and by DNA probes, a time-consuming method. This report describes a polymerase chain reaction (PCR) procedure for the detection of the stn (NAG-ST) and sto (O1-ST) gene sequences that is rapid and specific, allowing toxin gene molecular characterisation. A total of 34 V. cholerae and V. mimicus isolates was examined for ST and CT genes. The NAG-ST gene sequence was amplified in 13 of 22 non-O1/non-O139 V. cholerae and three of five V. mimicus strains. A new enterotoxin gene sequence pattern was found with MseI and TaqI restriction endonuclease PCR fragment digestion of two V. cholerae isolates, in addition to the pattern anticipated from the Genbank sequence, and found with the other ST+. These results show that ST-PCR detection is useful for the characterisation of V. cholerae and V. mimicus.

Effect of glucose on photodynamic action of methylene blue in Escherichia coli cells.

The results of this work show that the resistance of Escherichia coli cells to the photodynamic action of methylene blue is increased by the addition of glucose to the media in which they are grown. It is postulated that the increased resistance may be due to lowered retention of the dye by cells grown in the presence of glucose, leading to the diminution in DNA damage revealed in the alkaline sucrose gradients. The role of cyclic adenosine-monophosphate in the protective action of glucose is discussed.

Rectal antinociceptive properties of alverine citrate are linked to antagonism at the 5-HT1A receptor subtype.

Serotonin (5-HT) is considered as a major mediator causing hyperalgesia and is involved in inflammatory reactions and irritable bowel syndrome. Alverine citrate may possess visceral antinociceptive properties in a rat model of rectal distension-induced abdominal contractions. This study was designed to evaluate the pharmacological properties of alverine citrate in a rat model of rectal hyperalgesia induced by 5-HTP (5-HT precursor) and by a selective 5-HT1A agonist (8-OH-DPAT) and to compare this activity with a reference 5-HT1A antagonist (WAY 100635). At 4 h after their administration, 5-HTP and 8-OH-DPAT increased the number of abdominal contractions in response to rectal distension at the lowest volume of distension (0.4 mL). When injected intraperitoneally before 8-OH-DPAT and 5-HTP, WAY 100635 (1 mg kg(-1)) blocked their nociceptive effect, but also reduced the response to the highest volume of distension (1.6 mL). Similarly, when injected intraperitoneally, alverine citrate (20 mg kg(-1)) suppressed the effect of 5-HTP, but not that of 8-OH-DPAT. However, when injected intracerebroventricularly (75 microg/rat) alverine citrate reduced 8-OH-DPAT-induced enhancement of rectal distension-induced abdominal contractions. In-vitro binding studies revealed that alverine citrate had a high affinity for 5-HT1A receptors and a weak affinity for 5-HT3 and 5-HT4 subtypes. These results suggest that 5-HTP-induced rectal hypersensitivity involves 5-TH1A receptors and that alverine citrate acts as a selective antagonist at the 5-HT1A receptor subtype to block both 5-HTP and 8-OH-DPAT-induced rectal hypersensitivity.

Influence of trimebutine on inflammation- and stress-induced hyperalgesia to rectal distension in rats.

The effects of trimebutine and its major metabolite, N-desmethyltrimebutine on inflammation- and stress-induced rectal hyperalgesia have been evaluated in rats fitted with electrodes implanted in the longitudinal striated muscle of the abdomen. Intermittent rectal distension was performed before and 3 days after induction of rectal inflammation by local infusion of trinitrobenzenesulphonic acid (in ethanol). Stress consisted of 2h partial restraint and rectal distension was performed before and 30min after the end of the partial restraint session. The animals were treated intraperitoneally with trimebutine or desmethyltrimebutine (5, 10 or 20mgkg(-1)) or vehicle 15min before rectal distension. Naloxone (1mgkg(-1)) or saline was injected subcutaneously before trimebutine and desmethyltrimebutine. Before treatment trimebutine at the highest dose (20mgkg(-1)) reduced the abdominal response to rectal distension for the highest volume of distension (1.6mL) whereas desmethyltrimebutine was inactive. After rectocolitis the abdominal response to rectal distension was enhanced and trimebutine at 5mgkg(-1) reduced and at 10 mgkg(-1) suppressed inflammation-induced hyperalgesia, an effect reversed by naloxone. Desmethyltrimebutine was inactive. Stress-induced hypersensitivity was attenuated or suppressed, or both, by trimebutine and desmethyltrimebutine at doses of 5, 10 or 20mgkg(-l); greater efficacy was observed for desmethyltrimebutine and the effects were not reversed by naloxone. It was concluded that trimebutine and desmethyltrimebutine are active against inflammation- and stress-induced rectal hyperalgesia but act differently. The effect of trimebutine on inflammation-induced hyperalgesia is mediated through opioid receptors.

Clinical approach to circumvention of multidrug resistance in refractory leukemic patients: association of cyclosporin A with etoposide.

Alternative therapy for refractory leukemic patients is being increasingly adopted. Circumvention of multidrug resistance represents a strategy that has been taken into account when conventional chemotherapy failed. In this work a group of 15 refractory, heavily pretreated, patients was enrolled in a circumvention protocol including etoposide (ETO) and cyclosporin A (CSA). All patients received etoposide prior to this schedule. Toxicity to circumvention protocol was acceptable and only one serious side-effect was observed. Two hematological clinical responses were seen, both of which were positive to P-glycoprotein immunostaining and exhibited in vitro modulation by CSA in cultures using the thymidine incorporation assay. Three out of four patients negative for P-glycoprotein achieved a minor response. Three out of six clinical failures were also negative for Pgp immunostaining one of which exhibited sinergistic effect between ETO and CSA. Our study suggests that hematological response to ETO and CSA association can be obtained in intensely pretreated leukemic patients. Several factors may affect the response such as clinical status before this therapy. Additionally, it also suggests that not all CSA effects on the combination ETO-CSA can be attributed to Pgp modulation.

Reduction of pancreatic enzyme content and mortality in experimental acute pancreatitis in rats.

A previous report has shown that undernutrition reduces the mortality of acute experimental pancreatitis probably by decreasing pancreatic enzyme content. Cerulein in physiological doses reduces the enzyme content of the pancreas without any harmful effect on the organ. The aim of the present study was to asses the effect of acute reduction of pancreatic enzyme content on the outcome of acute pancreatitis. Two groups of male Wistar rats weighing 230-250 g were studied: group I, 12-h fasted animals, and group II, ad libitum-fed animals who received cerulein at the inframaximal dose (0.2 microgram kg-1 h-1) for 2 h. Cerulein administration resulted in the reduction of the pancreatic contents of chymotrypsinogen (71%), trypsinogen (55%), proelastase (60%), amylase (62%) and cathepsin B (45%) (P < 0.05). However, no significant reduction in pancreatic phospholipase content was observed. Acute pancreatitis was induced in group I after 12-h fasting and in group II at the end of cerulein infusion by retrograde injection o 0.5 ml of 2.5% Na+ taurocholate into the pancreatic duct. Ascites volume and the degree of histologically observed lesions were similar in both groups, but 72-h mortality was 56% in the control group (10/ 18) and 23% (5/22) in the cerulein group (P < 0.05). We speculate that the reduction of pancreatic enzyme content may exert its beneficial effect in acute pancreatitis by decreasing the quantity of pancreatic enzymes reaching the circulation and consequently their pathogenic effects.

Hepatic damage during acute pancreatitis in the rat.

We studied the alterations in the metabolism of liver mitochondria in rats with acute pancreatitis. Male Wistar rats were allocated to a control group (group I) and to five other groups corresponding to 2, 4, 12, 24 and 48 h after the induction of acute pancreatitis by the injection of 5% sodium taurocholate into the pancreatic duct. Sham-operated animals were submitted to the same surgical steps except for the induction of acute pancreatitis. Mitochondrial oxidation and phosphorylation were measured polarographically by determining oxygen consumption without ADP (basal respiration, state 4) and in the presence of ADP (activated respiration, state 3). Serum amylase, transaminases (ALT and AST) and protein were also determined. Ascitic fluid, contents of amylase, trypsin and total protein were also determined and arterial blood pressure was measured in all groups. In ascitic fluid, trypsin and amylase increased reaching a maximum at 2 and 4 h, respectively. Serum amylase increased at 2 h reaching a maximum at 4 h. Serum transaminase levels increased at 12 and 24 h. After 2 h (and also 4 h) there was an increase in state 4 respiration (45.65 +/- 1.79 vs 28.96 +/- 1.50) and a decrease in respiration control rate (3.53 +/- 0.09 vs 4.45 +/- 0.08) and in the ADP/O ratio (1.77 +/- 0.02 vs 1.91 +/- 0.01) compared to controls (P < 0.05). These results indicate a disruption of mitochondrial function, which recovered after 12 h. In the 48-h groups there was mitochondrial damage similar to that occurring in ischemic lesion. Beat-to-beat analysis (30 min) showed that arterial blood pressure remained normal up to 24 h (111 +/- 3 mmHg) while a significant decrease occurred in the 48-h group (91 +/- 4 mmHg). These data suggest biphasic damage in mitochondrial function in acute pancreatitis: an initial uncoupled phase, possibly secondary to enzyme activity, followed by a temporary recovery and then a late and final dysfunction, associated with arterial hypotension, possibly related to ischemic damage.

"Red complex" (Bacteroides forsythus, Porphyromonas gingivalis, and Treponema denticola) in endodontic infections: a molecular approach.

OBJECTIVE: The "red complex," composed of Bacteroides forsythus, Porphyromonas gingivalis, and Treponema denticola, is implicated in severe forms of periodontal diseases. The purpose of this study was to assess the occurrence of the red complex in root canal infections through the use of a sensitive technique-the 16S rDNA-directed polymerase chain reaction (PCR). STUDY DESIGN: Samples were obtained from 50 necrotic pulps with periradicular pathosis. Ten cases were diagnosed as acute periradicular abscesses. DNA was extracted from the samples and analyzed with a PCR-based identification assay. RESULTS: At least 1 member of the red complex was found in 33 of 50 cases. T denticola, P gingivalis, and B forsythus were detected in 44%, 30%, and 26% of the cases, respectively. The red complex was found in 4 of 50 cases. No particular signs or symptoms were associated with the presence of these bacterial species. CONCLUSIONS: Despite what is indicated in reports with respect to marginal periodontitis, red complex bacteria-either singularly or collectively-was not associated with any particular pattern of clinical symptoms. However, because the bacterial species from the red complex are recognized oral pathogens, their occurrence in root canal infections suggests that they may play a role in the pathogenesis of periradicular diseases.

[Protective effect of reduction of pancreatic enzyme content on the pulmonary disease induced by acute pancreatitis]. / Efeito protetor da redução do conteúdo enzimático do pâncreas na lesão pulmonar da pancreatite aguda.

Lung injury develop in up to 50-70 percent of patients with acute pancreatitis. Cerulein in physiological doses reduces the rate mortality of pancreatitis by decreasing the enzyme content of the pancreas. In order to assess the effect of acute reduction of pancreatic enzyme content on the pancreatitis pulmonary injury, pancreatitis was induced in Wistar rats by intraductal injection of 5 per cent sodium taurocholate: group I, with pancreatitis; group II, pancreatitis after decreasing pancreatic enzyme content; group III, control group. Dye Evans blue was used to evaluate the lung injury. The pancreatitis pulmonary injury was smaller in group II than group I (P < 0.05) but it was similar to control group (group III). In conclusion, it is speculated that the reduction of the pancreatic enzyme content reduces the pancreatitis pulmonary injury by decreasing the quantity of pancreatic enzymes reaching the systemic circulation.

[Essential arterial hypertension: psychopathology, compliance, and quality of life]. / Hipertensão arterial essencial: psicopatologia, compliance e qualidade de vida.

OBJECTIVE: To evaluate the influence of psychological and psychopathological factors and quality of life on hypertension, its treatment and patient compliance. DESIGN: Case-control study. SETTING: Primary Health Care Center in Oporto. PATIENTS OR PARTICIPANTS: Forty nine patients (pts) with essential hypertension (HT), 35 female and 14 male, mean ages: 52 +/- 11 yrs and 59 +/- 10 yrs, respectively, and 39 normotensive controls (NT)--18 female and 21 male, mean ages: 37 +/- 15 yrs and 42 +/- 15 yrs, respectively, were recruited from the same General Practice. METHODS: Hypertension was classified according to the Joint National Committee criteria. The following psychometric evaluations were used: the Beck Depression Inventory, the Hopkins Symptom Distress Checklist, the Psychological General Well-Being Schedule and the Eysenck Personality Inventory. RESULTS: 1. The hypertensive pts differed from the normotensive pts as they scored significantly higher in somatization (p = 0.07), aggression/hostility (p = 0.036), index of psychological distress (p = 0.08) and, neuroticism (p = 0.09); 2. the hypertensive pts showed lower scores of quality of life (p = 0.019); 3. the biochemical parameters studied (uric acid, urea, creatinine, glicose, total cholesterol, HDL cholesterol, trigliceride, transaminases and gammaglutamil transferase) did not show statistically significant correlation with the psychological variables and quality of life items studied; 4. the electrocardiographic and echocardiographic alterations, corresponding to the severity of the clinical situation, were not associated with statistically significant differences in depression and quality of life; 5. the angiotensin converting enzyme inhibitor, the calcium antagonist and the beta-blockers showed no statistically significant influence on the psychological scores studied. However, pts receiving diuretics showed higher scores of somatization (p = 0.0008) and obsession/ compulsion (p = 0.02) and, lower scores of quality of life (p = 0.04); 6. a better compliance was associated with better psychological scores, and somatization scored with statistical significance (p = 0.03). CONCLUSIONS: The hypertensive pts differed from the normotensive pts as they scored significantly higher in aggression/hostility and lower in quality of life. No statistically significant differences were found among the psychological variables in the pts with cardiac involvement. A better compliance was associated with better psychological scores. The results of this study lead us to suggest that when treating pts with HT, the most appropriate therapeutic attitude should attempt to avoid both therapeutic withdrawal and lack of medical control.

[The new biomedical and health research program of the European Economic Community--BIOMED 1 (1990-94)]. / O novo programa de investigação biomédica e de saúde da CEE--BIOMED 1 (1990-94).

The new EEC Biomedical and Health Research Programme (BIOMED 1) is presented. After a brief introduction on the background of the Programme--in which an overview of the ongoing Medical and Health Research Programme (MHR 4) is given--the main features of the new Programme are pointed out. On what concerns its aims and objectives, concertation and coordination of national research programmes at community level are stressed as well as the need to apply to the maximum possible extent the principles of subsidiary and community added value, in order to ensure an European dimension to the actions which are going to be supported and to achieve the harmonization of research methodologies, procedures and data. As regards the content of the Programme a description is made of its four main areas: Area 1--Development of co-ordinated research on prevention, care and health systems (main topics: drugs and the administration of medicines; risk factors and occupational health; biomedical technology; health services research); Area 2--Major health problems and diseases of great socio-economic impact (AIDS; cancer; cardiovascular disease; mental illness and neurological disease; the ageing process, and age-related health problems and handicaps); Area 3--Human Genoma Analysis (improvement of the genetic map; physical mapping; DNA sequencing; data-handling and databases; technology development and applications of human genome analysis); Area 4--Research on biomedical ethics (compilation of legislation; evaluation of questions of biomedical ethics linked with the Programme; evaluation of the social impact of the Programme and its risks).(ABSTRACT TRUNCATED AT 250 WORDS)

[The effect of the woman's age, the rate of cleavage and embryo quality on obtaining a pregnancy by in-vitro fertilization]. / Repercussão da idade da mulher, da taxa de clivagem e da qualidade embrionária, na obtenção de gravidez por fertilização in vitro.

Multiple factors influence the probability of obtaining a pregnancy through in vitro fertilization (IVF) and embryo transfer (ET). This retrospective study was designed to assess their importance in order to improve prognostic ability and treatment success. 341 consecutive embryo transfer cycles using the same ovarian stimulation protocol were considered and divided in two main groups: 92 cycles in which a clinical pregnancy was achieved and 249 cycles without success. All the embryo transfers were performed in patients from the in vitro fertilization program of the Human Reproductive Unit, Santa Maria Hospital, Lisbon, between January 1991 and December 1993. No significant differences were found between the two groups studied concerning the IVF indications, ovarian response to the stimulation, sperm quality, oocyte maturation and mean number of oocytes retrieved per patient. The women's age was higher in the group which did not achieve a pregnancy, when compared with the pregnant group (p < 0.001), showing a decline of success after the age of 35. Overall oocyte fertilization rate was 88.2% in cycles with pregnancy and 83.5% in cycles without pregnancy (p < 0.02). In the pregnant patients, there was a significantly higher rate of embryo transfers in which all the embryos received had reached at least the four-cell stage at 42-43 hr postinsemination, compared with the non pregnant patients (82% versus 63%, p < 0.001). All the 92 pregnancies originated from transfers of at least one embryo that had undergone two or more mitotic divisions.(ABSTRACT TRUNCATED AT 250 WORDS)