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The adenomatous polyposis coli (APC) protein is a negative regulator of the mitogenic transcription factor beta-catenin by stimulating its proteasomal degradation. This involves several APC domains, including the binding sites for axin/conductin, the recently described beta-Catenin Inhibitory Domain (CID) and the third 20 amino acid repeat (20R3) that is a beta-catenin-binding site. The four 15 amino acid repeats (15R) and the 20R1 are also beta-catenin-binding sites, but their role in beta-catenin degradation has remained unclear. We show here that binding of beta-catenin to the 15R of APC is necessary and sufficient to target beta-catenin for degradation whereas binding to the 20R1 is neither necessary nor sufficient. The first 15R displays the highest affinity for beta-catenin in the 15R-20R1 module. Biallelic mutations of the APC gene lead tocolon cancer in familial adenomatous polyposis coli (FAP) and result in the synthesis of truncated products lacking domains involved in beta-catenin degradation but still having a minimal length. The analysis of the distribution of truncating mutations along the APC sequence in colorectal tumours from FAP patients revealed that the first 15R is one target of the positive selection of mutations that lead to tumour development.

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The effect of sulfate in drinking water at concentrations of 600, 1,200, and 1,800 mg/L on nursery pig performance and health was evaluated over 28 days on 415 weaned pigs. Sodium sulfate and magnesium sulfate were evaluated in combination at concentrations of 600, 1,200, and 1,800 mg/L, and independently at concentrations of 600 and 1,800 mg/L in the drinking water. Seven treatment groups and 1 control group were evaluated for mean gain, feed consumption, water consumption, feed conversion, prevalence of diarrhea, and evidence of common post-weaning enteric pathogens. Statistical analysis was performed, using analysis of variance with repeated measures including initial pig weight as a covariate. Prevalence of diarrhea was analyzed nonparametrically with a repeated measures design. Results indicated that pigs drinking 600, 1,200, or 1,800 mg of sulfate/L water had increased prevalence of nonpathogenic diarrhea during the trial period. There was a trend for increased water consumption corresponding to increased sulfate in the water. Differences in mean daily gain, feed consumption, or feed-to-gain ratios were not observed. Forty-five pigs were treated at least once during the trial and 4 pigs died, resulting in a nursery morbidity of 11% and mortality of 0.96%. Fourteen isolates of enterotoxigenic Escherichia coli were found and rotavirus was isolated from 1 pig. Pigs in this study were not exposed to transmissible gastroenteritis virus. Except for an increase in fecal moisture content (not associated with pathogenic diarrhea), concentrations of up to 1,800 mg of sodium, magnesium, or a combination of sodium and magnesium sulfate/L had no adverse effect on nursery pig performance.

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Faeces and, or, paired sera were collected from cows in six dairy herds with classical winter dysentery. Similar samples were collected from cows in three other dairy herds experiencing non-haemorrhagic diarrhoea during the survey period. Coronavirus was the only enteric pathogen identified by immune electron microscopy (IEM) in all six outbreaks, occurring in 26 of 29 (90 per cent) of the affected cows and in one of 11 normal cows from the same herds. Nineteen of 26 affected cows (73 per cent) developed greater than four-fold increases in neutralising antibody titres to the Mebus strain of bovine coronavirus, compared with two of eight normal cows in the same herds. No cows showed greater than four-fold increases in antibody titres to bovine virus diarrhoea virus. None of the cows from the three herds with non-haemorrhagic diarrhoea shed coronavirus in faeces detectable by IEM or developed greater than two-fold rises in coronavirus antibody titres in paired sera. No enteric pathogens were identified in two of the herds. However, two cows in the third herd shed a group B rotavirus detected by IEM. These findings provide additional evidence for a possible role for bovine coronavirus in the aetiology of winter dysentery. Furthermore, this is the first report of a group B rotavirus associated with diarrhoea in adult cattle.

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A cell culture immunofluorescence (CCIF) assay was optimized for detection of porcine pararotavirus (group C rotavirus) in intestinal contents. The greatest viral infectivity was observed when MA104 cells (5 days after subculturing) were rinsed and refed in serum-free medium before inoculation, pancreatin was added to the inocula, and the inocula were centrifuged onto the cells. Gentamicin treatment of pararotavirus samples to reduce bacterial contamination also reduced the viral infectivity of these samples for MA104 cells. An indirect CCIF assay was used to determine the prevalence of pararotavirus and rotavirus antibodies in pig sera. In pigs from four herds, pararotavirus antibodies were detected in 100% (68 of 68) of adults and 59% (24 of 41) of weanling pigs, while 86% (24 of 28) of nursing pigs from 12 herds had pararotavirus antibodies. The physicochemical properties of pararotavirus were examined and compared with those of group A rotaviruses by using the CCIF assay to quantitate in vitro changes in viral infectivity. Pararotavirus was inactivated (greater than or equal to 99% reduction in titer) by heating to 56 degrees C for 30 min, was slightly labile at pH 3 (16 to 34% reduction in titer), and was stable at pH 5 (0 to 17% reduction in titer) and in either (3 to 19% reduction in titer). One group A rotavirus (Gottfried strain) was stable at 56 degrees C (0% reduction in titer), whereas the OSU strain of group A rotavirus was inactivated at this temperature (99% reduction in titer).

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The onset of fecal shedding, serogroup involvement, and association of hemolytic Escherichia coli with the postweaning diarrhea syndrome were studied in swine. The only E coli O antigen, detected by slide agglutination with the antisera used, was O157; K antigens were not detected. The ligated intestinal loop test (LILT) was used for enterotoxigenicity testing. All O157 serogroup isolates (n = 9) were hemolytic, and 89% (8 of 9) were LILT positive. Of all hemolytic isolates tested, 59% (10 of 17) were LILT positive. Twenty-seven nonhemolytic isolates were tested for enterotoxigenicity; of these, 45% (12) were LILT positive. The onset of postweaning diarrhea coincided with the modal onset of hemolytic E coli shedding at postweaning day 7. At postweaning day 7, hemolytic E coli shedding was concurrently associated with diarrhea (P less than 0.0005), whereas later during the postweaning period, this was not the situation.

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A commercially available, porcine-origin rotavirus vaccine was evaluated for efficacy against postweaning diarrhea due to rotavirus infection in pigs. Weight gains were compared at 5 intervals after weaning. Visual scoring was used to evaluate fecal consistency. Rectal swab specimens were cultured for hemolytic E coli and evaluated for rotaviral antigen by use of enzyme-linked immunosorbent assay. Milk from dams and sera from pigs and dams were evaluated for rotavirus-neutralizing antibodies by use of a plaque-reduction test. Significant differences between vaccinates and controls were not found in the determinants evaluated. Selected rotavirus-positive fecal and rectal swab specimens were examined for double-stranded (ds) RNA by use of direct electropherotyping, and the results were compared with the dsRNA pattern of rotavirus propagated from the vaccine. Only electropherotypes typical of field strain virus were found in the fecal and rectal swab specimens evaluated. Sera from guinea pigs and from a gnotobiotic pig immunized against the field strain rotavirus neutralized Ohio State University strain rotavirus (homologous to the vaccine rotavirus strain), but did not neutralize the Gottfried strain of rotavirus. This indicated that, although the dsRNA electropherotypes of the field and vaccine strains of the virus were different, the serotypes were similar, if not identical.

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A procedure for extracting rotaviral double-stranded ribonucleic acid (RNA) directly from fecal and intestinal specimens collected from calves and pigs is described. This procedure provides a rapid, simple, reproducible method of obtaining rotaviral double-stranded RNA preparations suitable for electrophoretic analysis in polyacrylamide-agarose composite gels. The rotaviral genome electrophoretic migration pattern produced by double-stranded RNA extracted directly from a specimen by this procedure was qualitatively identical to the electrophoretic migration pattern obtained with double-stranded RNA extracted from purified rotavirus derived from the same specimen. Direct extraction of specimens containing porcine rotavirus-like virus by this procedure gave preparations that had electrophoretic migration patterns similar, but not identical, to the characteristic electrophoretic migration pattern of the rotaviral genome. Sufficient rotaviral double-stranded RNA could be extracted from 6 ml of fecal or intestinal specimen by this procedure to permit 15 or more electrophoretic assays.

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Enterotoxigenic Escherichia coli (ETEC) that were isolated from neonatal pigs and that did not react in preliminary tests for pilus antigen K88 were subjected to additional tests for K88 and for pilus antigens K99 and 987P. Four such isolates produced K88, 9 isolates produced K99, 55 isolates produced 987P, and the remaining 43 isolates produced none of the three pilus antigens (3P(-)). Immunofluorescence tests of ileal sections from pigs were more sensitive for 987P detection than was serum agglutination of bacteria grown from the ileum. Most ETEC that produced K88, K99, or 987P were enteropathogenic (adhered to ileal villi, colonized intensively, and caused profuse diarrhea) when given to neonatal pigs. In contrast, only 3 of the 43 ETEC that produced none of the pilus antigens were enteropathogenic. The isolates were also tested for the type of enterotoxin produced. The K88(+) isolates all produced heat-labile enterotoxin (LT) detectable in cultured adrenal cells (i.e., were LT(+)). None of the 987P(+), K99(+), or enterpathogenic 3P(-) isolates produced LT. However (except for a single K99(+) isolate), they all produced heat-stable enterotoxin detectable in infant mice (STa(+)). Sixteen isolates produced neither LT nor STa but did produce enterotoxin detectable in ligated intestinal loops of pigs (STb). Most of these LT(-) STa(-) STb(+) isolates were also K88(-), K99(-), and 987P(-) and non-enteropathogenic. One of them was K99(+) and enteropathogenic. Our conclusions are as follows. (i) Most enteropathogenic ETEC from neonatal pigs produce either K88, 987P, or K99; however, there are some that produce none of the three antigens. (ii) Immunofluorescence tests for pilus antigens produced in vivo are recommended for the diagnosis of ETEC infections. (iii) Reports of LT(-) STa(-) STb(+) swine ETEC are confirmed; furthermore, such isolates can be enteropathogenic.

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A 3-day-old suckling pig with diarrhea was necropsied, and immunofluorescent microscopic examination of the small intestinal mucosa, together with immune electron microscopic examination of the large intestinal contents, provided a presumptive diagnosis of a concurrent infection with transmissible gastroenteritis (TGE) virus and porcine rotavirus. Immunofluorescent microscopic, immune electron microscopic, and serologic data obtained from gnotobiotic pigs experimentally inoculated with the large intestinal contents of the suckling pig confirmed this diagnosis. Two gnotobiotic pigs, convalescent from previous TGE viral infections, became infected with porcine rotavirus only. However, another gnotobiotic pig, convalescent from a previous porcine rotaviral infection, became infected with TGE virus only, following inoculation with the large intestinal contents of the suckling pig.

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In general health examinations of 2,447 4-year-old children in a certain area of southern Sweden, comprising 95.1% of the total population of that age, 52 children (2.1%) were diagnosed as having minimal brain dysfunction. After 7--9 years the children were reexamined and their parents and teachers were interviewed. Although no specific treatment, like stimulant drugs, was given, the children were much improved as they grew older: their hyperactivity had diminished, their behavior did not cause as much trouble, and their remaining neurological disturbances were small. However, the children manifested more problems in elementary school than other children, both regarding behavior, learning, slight neurological disturbances and visual disorders. Thus, the small group of children with minimal brain dysfunction symptoms in preschool age seem to run a certain risk of having trouble in school, although the symptoms are less conspicious as they grow and mature.

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Porcine rotavirus was shown to infect gnotobiotic pigs and induce an acute enteric disease clinically characterized by diarrhea, anorexia, depression, and occasional vomition. Onset of clinical signs correlated closely with the appearance of lesions within the small intestinal mucosa, and recovery from infection was associated with the regeneration of normal, functional villous epithelium. Villous atrophy, especially in the caudal two-thirds of the small intestine, was the consistent lesion observed in pigs with clinical signs of rotaviral infection. Villi were often short, blunt, and covered with cuboidal epithelial cells. Immunofluorescent microscopy methods demonstrated that the principal site of rotaviral replication was the villous columnar epithelial cells in the small intestine.

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A rotavirus (reovirus-like agent) was associated with diarrheal diseases occurring in 1- to 4-week-old suckling pigs in 8 herds and in weaned pigs in 2 herds. Transmissible gastroenteritis virus was also detected in 2 of these herds, as was enteropathogenic Escherichia coli in 5 herds. Morbidity was generally greater than 80% in pigs of the affected age group within these herds, and mortality from diarrhea ranged from 7 to 20%. The disease due to rotavirus in suckling pigs appeared similar to the syndrome commonly referred to as milk scours, white scours, or 3-week scours. Diarrhea and villous atrophy, resembling that seen in transmissible gastroenteritis, occurred in naturally infected pigs and in gnotobiotic pigs experimentally infected with rotavirus. Diagnosis was accomplished by immune electron microscopy of intestinal contents and by immunofluorescent staining of enterocytes. A massive infection of enterocytes with rotavirus was demonstrated by immunofluorescence, which helps explain the pathogenesis of this disease. The apparent rarity of clinical rotaviral infections in suckling pigs greater than 7 days old is probably due to the acquisition of passive immunity from immune sows.

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Immune electron microscopy (IEM) was developed as a diagnostic aid for detecting and identifying transmissible gastroenteritis virus and rotavirus (reovirus-like agent) in fecal and intestinal contents from cases of gastroenteritis in young pigs. Variables involved in use of direct IEM and its sensitivity were determined. Aggregates of virus coated with specific antibody were seen in virus samples mixed with homologous convalescent antiserum, but not in control samples containing preexposure serum or antibody directed against a heterologous virus. At least a ten fold enhancement of the sensitivity of direct IEM for virus detection was accomplished using indirect IEM employing rabbit anti-porcine IgG to further aggregate virus-antibody complexes. The technique was used to investigate the size and morphology of the porcine rotavirus. Particles ranged from 55 to 70 nm in diameter and had capsomere structures. Morphologically, the porcine rotavirus resembled the calf and human rotaviruses. By IEM, employing specific antiserums for each virus, porcine rotavirus was found to be antigenically related to these 2 viruses, but not to the reovirus type 3.

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Pregnant swine were vaccinated with 1 of 2 enteropathogenic strains of Escherichia coli, and their pigs were challenge inoculated with the homologous strain 1.5 hours after the entire litter was born. Seventeen sows were vaccinated orally by feeding viable cultures on 3 consecutive days, 6 sows were given 2 intramuscular (IM) injections of viable cultures at 10- to 14-day intervals, 6 sows were given 2 IM injections fo formolized cultures at 10- to 14-day intervals, and 12 sows were not vaccinated. The pigs from sows which had been orally vaccinated with viable cultures were protected during the 10-day observation period against diarrhea, as well as against death, when the newborn pigs were challenge inoculated with the homologous strain. Most of the pigs from sows which had been vaccinated IM were protected against death, but few neonates were protected against the diarrheal effects of challenge exposure with the homologous strain. Challenge-inoculated pigs suckling nonvaccinated sows had diarrhea and became dehydrated, and many died. Fewer viable E coli were usually recovered from the homogenized intestinal contents or intestinal segments of newborn pigs which did not have diarrhea than from similar specimens of diarrheal pigs. Microscopic examination of segments of the small intestine revealed that large numbers of E coli were closely associated with the ileal mucosa of newborn pigs killed in the acute phase of neonatal enteric colibacillosis caused by either strain, but organisms were not detected in this location in the pigs which remained normal after challenge exposure. It is concluded that swine which have been vaccinated by feeding large numbers of viable E coli late in gestation can provide good protection to their suckling pigs against the effects of challenge inoculation with the homologous enteropathogenic strain.

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