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The aim of this study was to choose the optimal variant of PCR examination of faeces to detect Echinococcus multilocularis infection which would allow to reduce the influence of different inhibitors in faeces. The investigation was carried out by comparison of 3 different methods of DNA isolation from faeces and different DNA dilutions used in PCR. Thirty five intestines of red foxes were used. Small intestines were examined by the sedimentation and counting technique (SCT). Faeces were collected from the rectum for PCR and flotation. DNA were isolated with the use of 3 different methods. Two methods were dedicated for faeces: method 1 (M1) - for larger samples and method 2 (M2) - for standard samples. The third method, method 3 (M3), was not dedicated for faeces. DNA samples were tested by nested PCR in 6 variants: not diluted (1/1) and 5 diluted (1/2.5, 1/5, 1/10. 1/20, 1/40). E. multilocularis was found by SCT in 18 from 35 (51.4%) intestines. Taenia-type eggs were detected only in 20.0% of faecal samples. In PCR the highest number of positive results (45.7%) were obtained during examination of DNA isolated by M1 method, and then 40.0% and 34.3%, respectively, for M2 and M3. In some samples positive results in PCR were obtained only in diluted DNA. For example, 8 from 12 positive samples isolated by M3 method gave the PCR negative results in non-diluted DNA and positive only after dilution 1:2.5, 1:10 or 1:20. Also 3 samples isolated by methods dedicated for stool gave positive results only after DNA dilution. The investigation has revealed that in copro-PCR for detection of E. multilocularis infection additional using of diluted DNA (besides non diluted) can avoid false negative results causing by PCR inhibition. In the best method of DNA isolation (M1), the use of non diluted DNA sample together with diluted in proportion 1:10 seems to be optimal.
The aim of this investigation was to determine the prevalence of coccidian invasions in suckling piglets and sows in different regions of Poland. The research was conducted in 2003 in 12 out of 16 Polish provinces in 3 types of farms: large (>100 sows), medium (25 to 100 sows) and small (<25 sows). Diarrhea in unweaned piglets was observed in all the examined farms. Overall, 473 litters of suckling piglets from 65 farms and 237 mother sows were examined. Feces were examined by the modified McMaster method. Isopsora suis were found in 146 litters (30.9%) from 49 farms (75.4%). Coccidia from the genus Eimeria (E. polita, E. debliecki, E. suis) were found in only 16 litters (3.4%) from 8 farms (12.3%). In large farms, I. suis invasion was detected in 35.2% of the litters whereas Eimeria spp. in only 1.9 % of them. In medium farms, I. suis was found in 21.8% of the litters and Eimeria spp. were not detected. In small farms I. suis was found only in 14.7% of the litters whereas Eimeria spp. in as many as 29.4%. I. suis was found in 15 (6.3%) of the fecal samples of the sows. For Eimeria spp. (E. debliecki, E. polita, E. suis, E. perminuta) the sow incidence rate was also 6.3%. Moreover in only 2 sows (0.8%) both types of coccidia occurred together. From 61 sows fostering I. suis infected piglets, only 9 (14.8%) shed I. suis oocyst and as little as 4 (6.5%) shed Eimeria oocyst. The remaining 48 sows (78.7%) did not shed coccidian oocysts. The results of this investigation show the high prevalence of Isospora suis in suckling piglets in large pig farms in Poland.
The aim of investigations was to determine the stability of ELISA components preserved under different conditions. The following components for the ELISA were used: the secreted/excreted antigen prepared from H. bovis L1 larvae, its fraction containing hypodermine A, conjugat, positive and negative control sera. Freezing at -20°C, lyophilization and preservation at 4°C were applied. Seven kits for the ELISA containing components preserved using described methods, were prepared., The identical packets of sera were examined by the ELISA directly after preparation of the kits (day '0'), and next, at 3-4 week intervals for 9 months. The experiments showed that the best method of conservation of ELISA components for the detection of H. Bovis antibodies was lyophilization. The kits prepared in this way did not lose their characteristics for 9 months and their specificity and sensitivity were close 100%.
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