Typing of avian pathogenic Escherichia coli strains by REP-PCR

In the present study the repetitive extragenic palindromic (REP) polymerase chain reaction (PCR) technique was used to establish the clonal variability of 49 avian Escherichia coli (APEC) strains isolated from different outbreak cases of septicemia (n=24), swollen head syndrome (n=14) and omphalitis (n=11). Thirty commensal strains isolated from poultry with no signs of these illnesses were used as control strains. The purified DNA of these strains produced electrophoretic profiles ranging from 0 to 15 bands with molecular sizes varying from 100 bp to 6.1 kb, allowing the grouping of the 79 strains into a dendrogram containing 49 REP-types. Although REP-PCR showed good discriminating power it was not able to group the strains either into specific pathogenic classes or to differentiate between pathogenic and non-pathogenic strains. On the contrary, we recently demonstrated that other techniques such as ERIC-PCR and isoenzyme profiles are appropriate to discriminate between commensal and APEC strains and also to group these strains into specific pathogenic classes. In conclusion, REP-PCR seems to be a technique neither efficient nor universal for APEC strains discrimination. However, the population clonal structure obtained with the use of REP-PCR must not be ignored particularly if one takes into account that the APEC pathogenic mechanisms are not completely understood yet.

Postweaning multisystemic wasting syndrome in farmed wild boars (Sus scrofa) in Rio Grande do Sul

This report concerns an outbreak of postweaning multisystemic wasting syndrome in wild boars from commercial herds in Rio Grande do Sul, Brazil. The main clinical signs were progressive wasting, respiratory distress, diarrhea, and ocular discharges. Enhanced lymph nodes and spleens, ascites, hydrothorax, and non collapsed lungs were the principal gross lesions. Varying degrees of lymphohistiocytic infiltrates in lymph nodes, spleen, and intestines were the most significant microscopic changes. Although a total of 26 animals had showed clinical and pathological findings consistent with the disease, only in 11 of them it was possible to confirm a PMWS diagnosis through the association of specific lesions in lymphoid tissues with the presence of the viral antigen detected by immunohistochemistry. These findings were similar to those reported in the swine disease.

Transmission of porcine circovirus 2 (PCV2) by semen and viral distribution in different piglet tissues

Porcine circovirus infections are caused by the porcine circovirus 2 (PCV2). Among six different clinical manifestations involving respiratory, enteric, nervous and reproductive signs, the postweaning multisystemic wasting syndrome (PMWS) is the most important and studied disease. However, reproductive failures associated with PCV2 have been increasingly reported. Some studies have shown the possible contamination of sows by semen of PCV2 positive boars. In order to investigate the transmission of PCV2 by contaminated semen and its ability to infect the sow and piglets, 20 PCV2 negative sows were inseminated, 10 with negative boar semen and 10 with previously nested-PCR tested positive boar semen. The sows were weekly monitored and blood samples were collected. Based on the results, 4 out 20 sows were selected (1 sow was PCR negative and inseminated with a negative semen, 2 sows were PCR negative and inseminated with a positive semen and 1 sow was PCR negative and inseminated with a positive semen, but became PCR positive around the 30 days of pregnancy). After weaning, 12 male piglets, 3 of each sow, were selected and maintained under isolation. In order to investigate which organs harbored the virus, the young pigs were necropsied around 9 months of age. Samples of serum collected monthly were tested by immunocitochemistry (ICC), and all 12 pigs serum converted. Samples of lymphoid, systemic and reproductive organs were analyzed by nested-PCR and immunohistochemistry (IHC). Evaluation of the samples by nested-PCR, revealed that several tissues were positive in 10 of 12 pigs, mainly the lymph nodes, bone marrow and spleen. Various samples were positive by IHC in 8 of 12 piglets, being the lymph nodes, tonsils and bulbourethral glands the most frequently positive. Thus, the results of testing different samples, in the 3 tests (ICC, nested-PCR and IHC) were complementary. These results show that PCV2 transmission through semen to the sows and piglets may occur and may also represent a potential risk for the herd.

Intestinal lesions in pigs affected with postweaning multisystemic wasting syndrome

Samples of mesenteric lymph nodes and intestines from 79 unthrifty 3- to 5-month-old postweaning pigs, confirmed as naturally affected with postweaning multisystemic wasting syndrome (PMWS), were studied. Pigs originated from 12 farms in southern Brazil and were selected on the basis of clinical signs and/or gross lesions suggestive of enteric disorder. Lymphohistiocytic infiltrates of varying intensity were associated with anti-porcine circovirus type 2 (anti-PCV2) immunostaining (IS) in samples of intestines and mesenteric lymph nodes from all pigs. Although most findings were similar to those described in PCV2-associated enteritis, anti-PCV2 IS in association with depletion of the goblet cell mucin stores (24 pigs), diffuse ileal villous atrophy and fusion (18 pigs), and dilatation of the lymphatic vessels (11 pigs) combined or not with lymphangitis were also observed. PCV2 antigen was immunohistochemically demonstrated in the cytoplasm and nuclei from intralesional epithelial cells, histiocytes, and endothelial-like cells in intestinal tissues. Together these findings imply an association with PCV2. The presence of co-infections by Lawsonia intracellularis, Brachyspira spp., Mycobacterium spp., Salmonella spp., rotavirus, parvovirus, coronavirus and enteric calicivirus with PCV2 in the intestinal lesions was investigated.

Gastric ulcers in pigs affected with postweaning multisystemic wasting syndrome

Samples of gastric lymph nodes and the stomachs from 24 pigs selected from herds affected by postweaning multisystemic wasting syndrome (PMWS) and sudden death associated with gastric ulcers were studied. Pigs were selected on the basis of unthriftiness, decreased feed intake, and wasting. The stomachs were opened, inverted, and classified into 0-3 score according the severity of the gross lesions present in pars oesophagica (non-glandulargastric mucosa). Selected samples were processed for paraffin embedding and stained with hematoxylin and eosin. Immunohistochemistry using anti-PCV2 (porcine circovírus type 2) antibody, anti-Helicobacter pylori antibody and a wide-spectrum anti-cytokeratin antibody was performed. Gross changes in pars oesophagea were classified according to the severity of lesions as score 3, 2, and 1 in 8, 6, 5 stomachs respectivelly. Microscopically, hyperplastic lymphoid follicles, lymphohistiocytic inflammatory infiltrates and focci of necrosis in the gastric mucosa were common findings. Large amounts of PCV2 antigen were observed in the cytoplasm and nuclei from intralesional cells and debris from the gastric glandular mucosal zone; however, in the fundus, anti-PCV2 immunostaining was restricted to the surface mucosal cells and foveolar compartment. All gastric lymph nodes were positive for PCV2 antigen. Anti-H. pylori immunostaining was seen in eleven cases, mainly in the antrum, on the mucosal surface and foveolar compartment. The association of the anti-PCV2 immunostaining with the glandular mucus-producing cells suggests a role for PCV2 as an additional factor for the swine ulcer development.

Facial paralysis and vestibular syndrome in feedlot cattle in Argentina

This paper reports 6 outbreaks of neurological disease associated with paralysis of the facial and vestibulocochlear nerves caused by intracranial space occupying lesions in feedlot cattle. The clinical signs observed were characterized by head tilt, uni or bilateral drooping and paralysis of the ears, eyelid ptosis, keratoconjunctivitis, and different degrees of ataxia. Morbidity and mortality rates ranged from 1.1 to 50% and 0 to 1%, respectively. Gross lesions observed included yellow, thickened leptomeninges, and marked enlargement of the roots of cranial nerves VII (facial) and VIII (vestibulocochlear). Histopathologically, there was severe, chronic, granulomatous meningitis and, in one case, chronic, granulomatous neuritis of the VII and VIII cranial nerves. Attempts to identify bacterial, viral, or parasitic agents were unsuccessful. Based on the morphologic lesions, the clinical condition was diagnosed as facial paralysis and vestibular syndrome associated with space occupying lesions in the meninges and the cranial nerves VII and VIII. Feedlot is a practice of growing diffusion in our country and this is a first report of outbreaks of facial paralysis and vestibular disease associated with space occupying lesions in Argentina.

Different pathways in the larval development of the crab Ucides cordatus (Decapoda, Ocypodidae) and their relation with high mortality rates by the end of massive larvicultures

One of the most limiting factors affecting the larval rearing of Ucides cordatus in the laboratory is a period of high mortality, which usually occurs late in the course of the larviculture during the metamorphosis from the zoeal to the megalopal phase. The objective of the present research was to analyze the post-embryonic development of U. cordatus on an individual basis and, in particular, to search for patterns linking disturbances in the molting process to the high larval death rates observed in massive larvicultures. A total of 50 larvae were individually reared from hatching to metamorphosis into the megalopal phase under controlled conditions, fed a combination of microalgae and rotifers. The survivorship rate was 70% until zoea V. The 35 surviving zoea V larvae followed two different pathways. Eleven underwent metamorphosis directly to megalopa, eighteen molted to zoea VI and six died as zoea V. In the last molting event, only two zoea VI larvae reached the megalopal stage, while the remaining sixteen died. In further observation under microscope, 13 of the dead zoea VI showed characteristics of the pre-molt stage and pereiopods disproportionably large in relation to the carapace. The observed pattern resembles the Molt Death Syndrome (MDS) described for other decapod species, in which larvae die in the late pre-molt phase of the molting cycle. We suggest that U. cordatus larvae develop disturbances in the molting process similar to the MDS described for other species and that these disturbances are related to a more complex pathway involving the emergence of larval stage zoea VI.

Oxidized LDL and physical fitness in healthy young men: associations with body composition, smoking, metabolic syndrome and androgen status

Background: In the past, oxidized low density lipoprotein (ox-LDL) has been associated with an unbeneficial lipid profile. This atherogenic lipid profile increases the risk of atherosclerotic cardiovascular diseases. Physical fitness has substantial effect on serum lipoprotein concentration as well as body composition and humoral responses, however interrelationships between ox-LDL and physical fitness have not been widely examined in a nationally representative sample. Aims: This thesis evaluates how cardiorespiratory and muscular fitness associate with ox-LDL lipids and how the other known risk factors of atherosclerosis might alter these associations. Subjects and Methods: The study cohort consisted of 846 healthy young males (mean age 25.1, SD 4.6) who were gathered by voluntary nationwide recruitment. Each participant conducted a series of physical fitness tests (cardiorespiratory and muscular fitness) and answered a detailed questionnaire that included lifestyle habits (i.e. smoking and leisuretime physical activity). Venous blood samples including ox-LDL and serum lipids were also collected. Results: Higher levels of ox-LDL were found in overweight and obese men, however, high cardiorespiratory fitness seemed to protect the overweight from high levels of ox-LDL. Young men who smoked and had poor cardiorespiratory or muscular fitness possessed a higher concentration of ox-LDL lipids when compared to comparable levels of cardiorespiratory or muscular fitness non-smoking young men. Metabolic syndrome was associated with increased levels of ox-LDL and high levels of ox-LDL combined with poor cardiorespiratory and abdominal muscle fitness seems to predict metabolic syndrome in young men. Also, participants with poor cardiorespiratory fitness and low levels of testosterone had higher levels of ox-LDL when compared to participants with high cardiorespiratory fitness / low testosterone as well as those with poor cardiorespiratory fitness / high testosterone. Conclusions: Good cardiorespiratory and muscular fitness protects young men from increased levels of ox-LDL lipids. This association was discovered in young men who were categorized as being overweight, smokers, metabolic syndrome or with low levels of testosterone. Being fit seems to prevent higher levels of ox-LDL, even in young healthy

Fowl adenovirus Group I as a causal agent of inclusion body hepatitis/hydropericardium syndrome (IBH/HPS) outbreak in brazilian broiler flocks

Commercial broiler flocks from a farm located in the State of São Paulo, Brazil, presented diarrhea, depression, increased mortality and poor weight gain. Upon post-mortem examination, classical signs of Inclusion Body Hepatitis/Hydropericardium Syndrome (IBH/HPS) were observed, including enlarged pale yellow-colored livers and straw-colored liquid in the pericardial sac. In addition, gross lesions were also observed in the kidneys, pancreas, thymus, intestines and gallbladder. Samples of these organs were analyzed by PCR for the detection of the hexon gene of the Fowl Adenovirus (FAdVs) Group I. The results were positive for both flocks (A and B) assayed by PCR. The macroscopic lesions associated with the detection of FAdV Group I by PCR in several of these affected organs allowed for the identification of IBH/HPS. In fact, this is the first report in Brazil of IBH/HPS in broilers, which identifies FAdVs group I as a causal agent of the disease. These findings may contribute to the worldwide epidemiology of the adenovirus-mediated hepatitis/hydropericardium syndrome.

Association of Alport's syndrome with HLA-DR2 antigen in a group of unrelated patients

A few family studies have evaluated HLA antigens in Alport's syndrome; however, there are no large population studies. In the present report, we studied 40 unrelated white patients with Alport's syndrome seen at the Unit of Renal Transplantation, Faculty of Medicine of Ribeirão Preto, São Paulo, Brazil. HLA-A, -B, -DR and -DQ antigens were typed using a complement-dependent microlymphocytotoxicity assay. A control white population (N = 403) from the same geographical area was also typed for HLA antigens. Although the frequencies of HLA-A and -B antigens of patients were not statistically different from controls, the frequency of HLA-DR2 antigen observed in patients (65%) was significantly increased in relation to controls (26%; P<0.001). The relative risk and etiologic fraction for HLA-DR2 antigen were 5.2 and 0.525, respectively. Although few immunological abnormalities have been shown in Alport's syndrome, in this report we emphasize the association of HLA molecules and Alport's syndrome. Besides the well-known inherited molecular defects encoded by type IV collagen genes in Alport's syndrome, the major histocompatibility alleles may be in linkage disequilibrium with these defective collagen genes

The diversity of abnormal hormone receptors in adrenal Cushing's syndrome allows novel pharmacological therapies

Recent studies from several groups have indicated that abnormal or ectopic expression and function of adrenal receptors for various hormones may regulate cortisol production in ACTH-independent hypercortisolism. Gastric inhibitory polypeptide (GIP)-dependent Cushing's syndrome has been described in patients with either unilateral adenoma or bilateral macronodular adrenal hyperplasia; this syndrome results from the large adrenal overexpression of the GIP receptor without any activating mutation. We have conducted a systematic in vivo evaluation of patients with adrenal Cushing's syndrome in order to identify the presence of abnormal hormone receptors. In macronodular adrenal hyperplasia, we have identified, in addition to GIP-dependent Cushing's syndrome, other patients in whom cortisol production was regulated abnormally by vasopressin, ß-adrenergic receptor agonists, hCG/LH, or serotonin 5HT-4 receptor agonists. In patients with unilateral adrenal adenoma, the abnormal expression or function of GIP or vasopressin receptor has been found, but the presence of ectopic or abnormal hormone receptors appears to be less prevalent than in macronodular adrenal hyperplasia. The identification of the presence of an abnormal adrenal receptor offers the possibility of a new pharmacological approach to control hypercortisolism by suppressing the endogenous ligands or by using specific antagonists for the abnormal receptors.

Effect of antibodies to intercellular adhesion molecule type 1 on the protection of distant organs during reperfusion syndrome in rats

We investigated kidney and lung alterations caused by intercellular adhesion molecule type 1 (ICAM-1) blockade after ischemia and reperfusion of hind limb skeletal muscles. Rats were submitted to ligature of the infrarenal aorta for 6 h. The animals were randomized into three groups of 6 rats each: group I, sacrificed after ischemia; group II, reperfusion for 24 h, and group III, reperfusion for 24 h after receiving monoclonal anti-ICAM-1 antibodies. At the end of the experiment, blood samples were collected for creatinine, lactate dehydrogenase, creatine phosphokinase, potassium, pH and leukocyte counts. Samples were taken from the muscles of the hind limbs and from the kidneys and lungs for histological analysis and measurement of the neutrophil infiltrate by myeloperoxidase staining. The groups did not differ significantly with regard to the laboratory tests. There were no major histological alterations in the kidneys. An intense neutrophil infiltrate in the lungs, similar in all groups, was detected. Myeloperoxidase determination showed that after reperfusion there was significantly less retention of polymorphonuclear neutrophils in the muscles (352 ± 70 vs 1451 ± 235 × 10² neutrophils/mg; P<0.01) and in the kidneys (526 ± 89 vs 852 ± 73 × 10² neutrophils/mg; P<0.01) of the animals that received anti-ICAM-1 before perfusion compared to the group that did not. The use of anti-ICAM-1 antibodies in this experimental model minimized neutrophil influx, thus reducing the inflammatory process, in the muscles and kidneys after ischemia and reperfusion of the hind limbs.

Positive correlation of serum leptin with estradiol levels in patients with polycystic ovary syndrome

Patients with polycystic ovary syndrome (PCOS) usually are obese, insulin resistant and hyperinsulinemic. The known association between leptin, obesity andinsulin action suggests that leptin may have a role in PCOS but this has only been addressed peripherally. This study was designed to assess the relationship between serum leptin and the anthropometric, metabolic and endocrine variables of obese (body mass index, BMI ³30 kg/m²) and non-obese (BMI <30 kg/m²) PCOS patients. Twenty-eight PCOS patients and 24 control women subdivided into obese and non-obese groups were evaluated. Leptin, androgens, lipids, gonadotrophins and insulin-glucose response to the oral glucose tolerance test were measured by radioimmunoassay in all participants. The assays were done all in one time. The areas under the insulin curve (AUC-I) and the glycemia curve were calculated to identify patients with insulin resistance. Mean leptin levels were not significantly higher in patients with PCOS compared to the control group (21.2 ± 10.2 vs 27.3 ± 12.4 ng/ml). Leptin levels were found to be significantly higher in the obese subgroups both in patients with PCOS (26.9 ± 9.3 vs 14.1 ± 7.0 ng/ml) and in the control group (37.3 ± 15.5 vs 12.9 ± 5.8 ng/ml). The leptin of the PCOS group was correlated with BMI (r = 0.74; P < 0.0001) and estradiol (r = 0.48; P < 0.008) and tended to be correlated with the AUC-I (r = 0.36; P = 0.05). Of the parameters which showed a correlation with leptin in PCOS, only estradiol and probably insulinemia (AUC-I) did not show a significant correlation with BMI, suggesting that the other parameters were correlated with leptin due to their correlation with BMI. Estradiol correlated with leptin in PCOS patients regardless of their weight.

A calcineurin inhibitory protein overexpressed in Down's syndrome interacts with the product of a ubiquitously expressed transcript

The Down's syndrome candidate region 1 (DSCR1) protein, encoded by a gene located in the human chromosome 21, interacts with calcineurin and is overexpressed in Down's syndrome patients. As an approach to clarifying a putative function for this protein, in the present study we used the yeast two-hybrid system to identify DSCR1 partners. The two-hybrid system is a method that allows the identification of protein-protein interactions through reconstitution of the activity of the yeast GAL 4 transcriptional activator. The gene DSCR1 fused to the GAL 4 binding domain (BD) was used to screen a human fetal brain cDNA library cloned in fusion with the GAL 4 activation domain (AD). Three positive clones were found and sequence analysis revealed that all the plasmids coded for the ubiquitously expressed transcript (UXT). UXT, which is encoded in human Xp11, is a 157-amino acid protein present in both cytosol and nucleus of the cells. This positive interaction of DSCR1 and UXT was confirmed in vivo by mating the yeast strain AH109 (MATa)expressing AD-UXT with the strain Y187 (MATalpha) expressing BD-DSCR1, and in vitro by co-immunoprecipitation experiments. These results may help elucidate a new function for DSCR1 and its participation in Down's syndrome pathogenesis.

Effects of melatonin on the ovarian response to pinealectomy or continuous light in female rats: similarity with polycystic ovary syndrome

The current study was conducted to investigate the relationship between melatonin and chronic anovulation. Adult (3-4 months old) female Wistar rats were submitted to pinealectomy: group I: pinealectomized ovariectomized melatonin-treated (N = 10); group II: pinealectomized ovariectomized placebo-treated (N = 12); group III: pinealectomized light-treated placebo-treated(N = 10) or maintained under continuous light; group IV: maintained under continuous light, ovariectomized melatonin-treated (N = 22); group V: maintained under continuous light, ovariectomized placebo-treated (N = 10); group VI: maintained under continuous light placebo-treated (N = 10). In order to assess ovarian modifications, unilateral ovariectomy was performed during the fourth month in groups I, II, IV, V and the other ovary was removed after 8 months. Ovariectomy was performed in groups III and VI only after eight months. Melatonin (200 µg/100 g body weight) dissolved in 0.02 ml absolute ethanol was injected intramuscularly daily during the last 4 months into groups I and IV. The other groups were treated with placebo (NaCl). The ovarian cysts were analyzed and their area, perimeter and maximum diameter, as well as the thickness of the ovarian capsule were measured. Daily colpocytological smears were performed throughout the study. Persistent estrous condition and ovarian cysts were observed in all groups. In pinealectomized rats the ovarian and vaginal alterations disappeared at the end of the study and in rats maintained under continuous light the vaginal and ovarian polycystic aspect was reversed only in those treated with melatonin. We conclude that melatonin may act on the ovarian response reverting chronic anovulation induced by pinealectomy or continuous light.