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1.
A novel in vitro flow through technique was developed and evaluated for immunodiagnosis of cystic echinococcosis in sheep using hydatid specific non-cross reactive 8-kDa protein. The 8-kDa protein was prepared from hydatid cyst fluid by DEAE-Sepharose fast flow anion exchange chromatography. In this flow through technique, the 8-kDa antigen was coated on the nitrocellulose membrane of flow through device. Protein A colloidal gold was used as detector. The evaluation of the technique was performed by comparing 150 known positive hydatid serum and known negative serum collected from sheep. The test was shown to be high sensitivity and specificity that were closely correlated with those of EITB. Furthermore the immunofiltration-based assay is rapid (2 min) and easy to perform with no requirement of special skill, reagent and instrumentation. This suggests the flow through technique is an acceptance alternative to be used in clinical laboratories lacking specialized equipments as well as large scale screening of cystic echinococcosis both in the field with animal and human populations.  相似文献   

2.
Serum antibody responses in sheep naturally or experimentally infected with Echinococcus granulosus and/or other larval cestodes were examined using an enzyme-linked immunosorbent assay (ELISA) with various antigens prepared from sheep hydatid cyst fluid ( SHCF ). Serum donors included: sheep experimentally infected with E. granulosus and their age-matched non-infected controls; sheep experimentally infected with other helminth parasites; sheep naturally infected with E. granulosus both from Tasmania and the Australian mainland; sheep from Tasmania naturally infected with larval cestodes other than E. granulosus; and naturally reared sheep completely free from infection with larval cestodes. Attempts were made to eliminate serological reactions which were not specific for E. granulosus by using a series of antibody affinity chromatography steps to deplete crude SHCF antigen; these included adsorption with a monoclonal antibody, 3EgH 29-2, removal of host IgG using rabbit anti-sheep IgG antibody, and removal of antigens which bound non-specifically to normal sheep immunoglobulin. The final affinity-depleted antigen product was designated AD SHCF . Specific serological reactivity in infected sheep was very low. Affinity depletion of SHCF using 3EgH 29-2 did not appear to increase the specificity of serological diagnosis of E. granulosus infection when experimentally infected sheep were compared with their non-infected controls provided the latter were age-matched with experimental animals. The other affinity adsorption steps significantly reduced non-specific background binding to antigen by normal sheep serum. Despite this reduction in background in the ELISA, only low levels of antibody could be detected in naturally-infected sheep.(ABSTRACT TRUNCATED AT 250 WORDS)  相似文献   

3.
An ELISA was developed and tested for its ability to detect antibodies against Salmonella enteritidis in chickens. Various features of the ELISA were evaluated and optimized. The outer membrane protein antigens selected by use of the protein immunoblotting method made the assay specific and sensitive. The assay was evaluated in chickens experimentally infected with S enteritidis. Blood samples collected at weekly intervals after experimental infection with S enteritidis were analyzed by ELISA. Results of the ELISA were compared with those of conventional serum plate and microagglutination tests. The ELISA was more sensitive and specific in the detection of S enteritidis infection than the other 2 conventional tests.  相似文献   

4.
The sanitary and economic impact of cystic echinococcosis is serious in those countries where it becomes endemic. Ultrasonography is one technique that may be used to diagnose this disease in endemic areas. In parasitized sheep, hydatid cysts appear sonographically as a round hypoechoic structure. Twenty two sheep destined for slaughter were studied sonographically and imaging findings compared to post-mortem findings. Three sheep with hydatid cysts were identified. Eighty additional sheep not destined for slaughter were also studied. Echinococcus granulosus cysts were detected in three animals. Forty sheep from a non-endemic area had no hepatic cysts. The in vivo sonographic study of sheep provides a useful screening tool for echinococcosis.  相似文献   

5.
For the surveillance of trichinellosis, the digestion method is reliable but also labour intensive. The serological methods for the detection of Trichinella-specific antibodies using ELISA offer a sensitive and relatively specific alternative. For serological studies, sera or plasma from blood samples are the most common source of antibodies, but although the concentration of antibodies is approximately 10-fold lower, muscle fluid can be a good alternative particularly for testing of wildlife samples. In the present study, an indirect ELISA technique was evaluated on both sera and muscle fluids from experimentally infected foxes, pigs, and wild boars using both excretory/secretory (E/S) antigens and a synthetic glycan antigen, beta-tyvelose. Although the synthetic antigen appears to be less sensitive than the E/S antigens, Trichinella-specific IgG antibodies were detected in both serum samples and muscle fluid samples from pigs, wild boars and foxes infected at levels which would be important for food safety or represent a significant reservoir for further transmission.  相似文献   

6.
From 1991 to 1993, an investigation into the epidemiology of cystic echinococcosis (CE) was carried out in the Falkland Islands to evaluate the progress of the hydatid eradication campaign. The prevalence of CE in sheep was assessed using abattoir and farm slaughter data, and the exposure of dogs to the parasite was estimated using immunological techniques. A total of 59 466 sheep was examined at slaughter for E. granulosus and T. hydatigena cysts and the entire dog population of the Falkland Islands (n = 908) was examined by ELISA for the presence of specific serum antibodies to E. granulosus (IgG, IgA and IgE). In addition, a subsample of dogs (n = 464) was tested for the presence of E. granulosus antigens in faeces (copro-antigens). The prevalence of CE in sheep increased significantly during the period of the study from 0.11% in 1991 to 0.47% in 1993. Nineteen (2.1%) of 908 dog sera tested were seropositive, and eight dogs (1.7%) of 464 tested were positive in the copro-antigen assay. The combined use of abattoir surveillance, specific antibody and copro-antigen assay suggested that there were several locations in the Falkland Islands where the life cycle of E. granulosus may still perpetuate. Specific deficiencies in the eradication effort in those locations could be identified through follow-up questionnaires.  相似文献   

7.
Immunodiagnosis in sheep presents problems of sensitivity and specificity, limiting its applicability in surveillance systems. The objective of this study was to develop a sensitive, specific and accessible technique for diagnosing cystic echinococcosis in naturally infected sheep and to evaluate the validity of necropsy as a reference test. A total of 247 sheep were studied at slaughterhouses, confirming the parasitological diagnosis with histology. Serum was processed with enzyme immunoassay (EIA) using three antigen preparations: total hydatid liquid (LHT), purified fraction of LHT (S2B) and purified lipoprotein (B). Western Blot (WB) was used as a control. EIA proved effective for differentiating Echinococcus granulosus from larval stage of Taenia hydatigena and intestinal cestodes in all three antigen preparations. Serums from macroscopically negative sheep were reactive to EIA and positive with WB. In the whole flock, sensitivity was 89.2% for LHT, 80.0% for S2B and 86.4% for B. Sensitivity in lambs was 78.6% for LHT, 75.0% for S2B and 64.3% for B. Macroscopic diagnosis at the time of slaughter was found to have limitations as a reference test for immunodiagnosis of cystic equinococcosis in sheep, so it was necessary to include histology and WB as reference tests. LHT was the antigen preparation of greatest value and EIA proved to be a sensitive and specific technique, adequate for surveillance systems and for evaluating control programmes.  相似文献   

8.
Polysaccharide antigens were obtained from either the secretions produced during in vitro cultivation of Echinococcus granulosus protoscoleces or from mouse hydatid cyst membranes by phenol extraction. When either of these antigens was used in an enzyme-linked immunosorbent assay antibody activities were detected in sera from sheep infected 27 or more weeks earlier with at least 100 E granulosus eggs. These antibody responses were significantly higher (P less than 0.05) than those of sheep infected with Taenia hydatigena or T ovis and tested with the E granulosus antigens. Very high cross-reacting antibody responses in sera from sheep recently infected with T hydatigena were only detected with the protoscoleces secretions antigen. Neither antigen was sufficiently sensitive or specific for serodiagnostic use. However, when sera were first tested with one antigen and then with the other, and only sera that were positive in both tests were regarded as positive, the overall sensitivity and specificity of this two antigen method increased to about 80 per cent.  相似文献   

9.
Sheep from the areas of Fondouk-Jeddid, Bir Mchergua and El Fahs, located in the Northeast of Tunisia, were examined by ultrasonography between 2001 and 2004 in order to assess their infection with Echinococcus granulosus, the agent of hydatid disease, and to evaluate this method as an efficient aire for hydatid cysts. A total of 1039 sheep, aged between 1 and 14 years was examined. The highest prevalence was found in sheep aged more than 8 years. The least infected animals were aged between 1 and 2 years. All hydatid cysts detected by ultrasound were located in the liver. In all age-groups, the dead cysts were more numerous than viable cysts. Eighteen positive sheep were autopsied and a comparison between ultrasound and autopsy results was performed. The results showed a prevalence of about 40% for the three areas. Ultrasonography allowed the cysts, deep or superficial to localize in the central or left part in relation to the caudal vena cava of the animals. Consequently, all the cysts were not detected with this technique. This work shows that ultrasonography confirms the importance of ovine hydatid cyst in Tunisia and that its use as a mass screening approach for cystic echinococcosis in sheep could be helpful for the monitoring of this disease in a hydatid control program without great stress for the animals.  相似文献   

10.
Coprological confirmation of ovine fasciolosis in the field, prior to out breaks of the disease and/or strategic antifluke medication, seem to be of little consequence. Efforts are, therefore, being made to evolve a putative antigen specific to serodiagnostic test for early diagnosis during prepatency. In the present investigation, 28 kDa cysteine proteinase was used in ELI SA and Western blot to detect Fasciola gigantica antibodies and further Dipstick-ELISA was developed for field application, using known positive monospecific sera from experimentally infected sheep with 100 F. gigantica metacercariae. Isolation of 28 kDa cysteine proteinase was achieved from bubalian origin flukes. The specific antigen, recognised homologous antifluke antibodies by Western blot as early as 2nd week post-infection (wpi) with 100% sensitivity, in sera samples of sheep harbouring 38 flukes and by 10th wpi in sheep harbouring 3-8 flukes. All sheep were found positive for the infection when ELISA and/or Dipstick-ELISA was applied from 4th wpi. In pooled sera of infected sheep, these were positive during 4th wpi.  相似文献   

11.
In Sweden echinococcosis is uncommon in domestic animals. In reindeer in the most northern part of the country echinococcosis was found. Of 1453 pairs of lungs, 23 pairs (1.6 %) were infected with hydatid cysts. These were of two types: typical well-developed hydatid cysts, which were found in nine of the 23 infected lungs, and collapsed hydatid cysts, which were found in 13 of the lungs. In only one lung pair both types of cysts were seen.  相似文献   

12.
A specific enzyme-linked immunosorbent assay (ELISA) for detection of antibodies to the porcine pathogen Lawsonia intracellularis was developed and evaluated using sera from na?ve, naturally infected as well as experimentally infected pigs. On the basis of 37 serum samples collected from experimentally infected pigs and 62 serum samples from naturally infected pigs the sensitivity of the ELISA was calculated to 98.0%. The specificity of the test was 99.3%, calculated on the basis of 273 serum samples collected in six herds free of L. intracellularis after medicated eradication. The novel ELISA was a specific and sensitive method for detecting specific antibodies, and may be a good alternative to the existing serological tests for L. intracellularis. It may be usable for diagnosis of proliferative enteropathy and for determination of a herd's epidemiologic status.  相似文献   

13.
The effect of Fasciola hepatica parasite burden on the detection of excretory/secretory (E/S) antigens in sera and feces of experimentally infected sheep was evaluated using a double antibody-based capture enzyme-linked immunosorbent assay (ELISA). Four groups of five sheep each were used. The first three groups were infected with 50, 100 and 200 metacercariae of F. hepatica, and the fourth group remained as non-infected control. On the day of infection and weekly thereafter, serum and fecal samples were taken. ELISA detected F. hepatica E/S antigen levels in serum from the first week post-infection (wpi) and in fecal supernatant from the fourth wpi, which were significantly (p<0.05) higher than controls. F. hepatica eggs were not detected until after the eighth wpi. The correlation between absorbance of E/S antigens in serum with the fluke burden was 0.77 (p<0.0001) and in feces 0.76 (p<0.0001) at 12th wpi. The sensitivity of the assay to detect E/S antigens in serum was 86.6% and in feces 93.3%. It is concluded that the ELISA technique used in this study offers a diagnostic alternative for detecting early infections of F. hepatica in sheep.  相似文献   

14.
An enzyme-linked immunosorbent assay (ELISA) procedure was used to quantitate milk and serum antibodies (IgG) to Staphylococcus aureus alpha and beta toxins, and S. aureus 2-8 and Smith diffuse strain capsular antigens. Milk samples were collected on two occasions. A comparison was made between levels of milk antibodies specific for the two toxins and capsular antigens for 41 cows that were infected with S. aureus on both sampling dates, and 18 cows not S. aureus-infected on either date. Staphylococcus aureus-infected cows were grouped according to somatic cell counts. All groups of infected cows, regardless of somatic cell counts, had significantly higher milk antibody levels to alpha and beta toxins than did the non-infected cows (P less than .002). Serum samples taken for 13 infected and 4 non-infected cows also indicated that significant elevations in anti-alpha toxin and anti-beta toxin IgG were present in S. aureus-infected cows, compared to non-infected cows. A similar immune response was not seen to capsular antigens, however. No significant differences were present between the two groups of cows for either milk or serum antibodies to Smith diffuse strain capsular antigens. Milk antibodies to 2-8 capsule were significantly elevated only in infected cows with somatic cell counts greater than 10(6)/ml, compared to non-infected cows; no differences were present for serum antibodies to 2-8 capsule between infected and non-infected cows. These results indicate that significant increases in milk (and possibly serum) antibodies to alpha and beta toxins are present in cows with chronic staphylococcal mastitis, apparently resulting from a systemic immune response to these toxins. There does not appear to be a similar immune response to capsular antigens.  相似文献   

15.
Two serotypes of the anaerobic bacterium Dichelobacter nodosus were used to experimentally infect young sheep resulting in infectious pododermatitis or footrot characteristic of the natural disease in sheep. The specific serotypes of D. nodosus were reisolated from the feet and identified using immunofluorescent microscopy of hoof scrapings. Prior immunization of sheep with a commercially available bacterin containing whole cell preparations of ten strains of D. nodosus resulted in serum IgG reactive to a serotype of D. nodosus common to the vaccine. Immunization also produced serum IgG reactive to a serotype of D. nodosus not incorporated in the vaccine. A less severe infection occurred in the immunized sheep than in the controls regardless of the serotype of bacteria used to infect them. Clinical lameness and lesion severity were milder in sheep infected with the serotype of D. nodosus common to the vaccine. Western blot analysis of sera from convalescent sheep showed cross-reactive antibodies to nonfimbrial cell surface proteins, as well as bacterial lipopolysaccharide. Such cross-reactivity may explain the partial protection seen in animals infected with a serotype distinctive from the ones in the vaccine. Despite the historical emphasis of fimbrial immunogens in ovine footrot this study using a new model of experimental ovine footrot suggests other surface antigens may also be important in protective immunity.  相似文献   

16.
The aim of this study was to determine the fertility and viability of hydatid cysts in slaughtered sheep and cattle. Cysts were collected from the liver and lungs of 169 sheep and 171 cattle infected with Echinococcus granulosus when slaughtered in industrial abattoir in Sari, Iran, 2007–2008. Fertility was determined by the examination of cyst fluid for the presence of protoscoleces. The viability of the protoscoleces was assessed by staining with 0.1% aqueous eosin solution. The fertility rates of hepatic cyst of sheep and cattle were 47.1% and 1.4%, respectively and the fertility rates of pulmonary cyst of sheep and cattle were 39.4% and 8.1%. In the sheep, the fertility of cysts in the liver was higher than that in lungs, but in the cattle the fertility of cysts in lungs was higher than liver. The viability of protoscoleces of fertile cysts for sheep and cattle were about 76.9% and 82.5%, respectively. Based on the finding in the present study, effort should be made to control transmission of cystic echinococcosis by safe disposal of Echinococcus cysts such that dogs cannot have access to the cysts.  相似文献   

17.
In 2000 Guarnera et al. proposed using ELISA in canine faeces collected from the ground to detect dogs infected with Echinococcus granulosus, thus determining sheep farms with active transmission. The objective was to evaluate the prevalence of E. granulosus infection in sheep farms of the Patagonia. Sheep farms were randomly selected in the Provinces of Río Negro, Chubut, Neuquén, Santa Cruz and Tierra del Fuego (areas with control programs) and La Pampa (comparison area). From one to three samples of fecal matter were obtained for each sheep farm, which were processed by means of copro-ELISA test with confirmation of positive samples by copro-Western blot. A total of 1042 samples were obtained from 352 sheep farms, 26 (7.3%) proving positive. Of these 5 (6.3%) were from La Pampa, 9 (13.8%) from Neuquén, 4 (4.7%) from Río Negro, 2 (2.9%) from Chubut, 1 (5.9%) from Santa Cruz and 5 (13.9%) from Tierra del Fuego. The identification of parasitized dogs is an essential activity upon which rests the strategy of control and surveillance. Arecoline tests or coproantigen test with fecal matter obtained directly from the dog contribute information on individual prevalence, while the use of coproantigens detected in ground-collected samples transfers the dog unit of observation to units of greater epidemiological value. In the present experience, the technique employed seems promising for its application in systems of epidemiological surveillance of cystic echinococcosis and in drawing a baseline on which to measure the progress of control programs in the Argentine Patagonia in subsequent years.  相似文献   

18.
A commercially available ELISA for detecting antibodies to liver fluke was evaluated for use in Australia. Milk and serum samples from cattle and sheep in which infection with Fasciola hepatica was confirmed by detection of eggs in faeces were used to estimate sensitivity. Similar samples collected from cattle and sheep outside the F. hepatica-endemic area were used to estimate specificity. The ELISA was also evaluated for detecting antibodies to F. hepatica in milk from sheep and antibodies to Fasciola gigantica in sera from cattle and buffaloes, but with small numbers of samples. In cattle, the sensitivity and specificity of the ELISA were 98.2% and 98.3% using serum and 97.7% and 99.3% using milk. In infected herds, 41.4% and 41.5% of animals were positive in the serum and milk ELISAs, respectively, whereas F. hepatica eggs were found in faecal samples from 26.5% of animals. In sheep, the sensitivity of the ELISA was 96.9% and the specificity was 99.4%. In infected flocks, 60.2% of animals were positive in the serum ELISA and F. hepatica eggs were found in faecal samples 52.2% of animals. There was perfect agreement in the ELISA between paired serum and milk samples collected from ewes. The assay detected antibodies in sera from cattle and buffaloes with natural and experimental F. gigantica infections. In the experimentally infected animals, antibodies were detected 2 weeks post-infection. We conclude that the ELISA will be a valuable tool for diagnosing F. hepatica infections in cattle and sheep. The assay may also be useful for diagnosing F. gigantica infections but further studies are required to establish sensitivity and specificity.  相似文献   

19.
A maedi-visna virus CA-TM fusion protein ELISA (MVV ELISA) was evaluated for the detection of antibody in sheep infected with North American ovine lentivirus (OvLV). The results of the MVV ELISA were compared with other assays for OvLV antibody and with viral infection in an intensively studied group of 38 sheep with a high prevalence of OvLV infection and disease. The sensitivity, specificity, and concordance of assays for OvLV antibody (MVV ELISA, indirect ELISA, Western blot, and AGID), virus (virus isolation, PCR, antigen ELISA), and OvLV-induced disease in each animal were compared with OvLV infection status as defined by a positive result in two or more of the assays. Five sheep met the criteria for absence of OvLV infection. The sensitivity of the MVV ELISA in detecting OvLV infected sheep was 64%, whereas the sensitivity of the other three tests for antibody ranged from 85 to 94%. All the antibody assays were 100% specific in this group of animals. Of the assays for virus, the PCR test had the highest sensitivity and the best concordance with OvLV infection, but it also had the lowest specificity of any of the virus or antibody assays. Among the antibody tests, the concordance of the MVV ELISA compared most favorably with the AGID test for detecting OvLV-infected sheep. Analysis of serum samples from 28 lambs experimentally-infected with one of three North American strains of OvLV suggested that there were no significant strain differences detectable by antibody assay. Twenty virus-inoculated lambs were positive by both the MVV ELISA and the AGID test, five lambs were MVV ELISA negative and AGID test positive, and three lambs were MVV ELISA positive and AGID test negative. No pre-inoculation samples were positive by either assay. In a longitudinal study involving seven lambs, antibodies to OvLV were detected by AGID 3-5 weeks post-inoculation, but were not detected by MVV ELISA until 5-10 weeks post-inoculation. Among 128 naturally and experimentally-infected sheep that were seropositive in the AGID test, the overall sensitivity of the MVV ELISA was higher in the naturally infected sheep (84%) than in the experimentally infected sheep (69%). The data indicated that the MVV ELISA represents a less sensitive, but specific alternative for the detection of OvLV antibodies.  相似文献   

20.
A survey of cystic echinococcosis (CE) in the water buffalo (Bubalus bubalis) of the Italian Mediterranean breed was carried out in Campania, a region of southern Italy. In addition, a molecular study was performed on 48 hydatid cysts coming from 48 water buffaloes in order to determine the Echinococcus granulosus strain(s) present in this host. Out of a total of 722 water buffaloes examined for CE, 76 (10.5%) were found infected. The average number of cysts per buffalo was 4.3 (minimum 1, maximum 45). Seventeen buffaloes had hydatid cysts only in the liver (with an average of 5 cysts/liver), 34 only in the lungs (with an average of 1.8 cysts/lungs), and 25 buffaloes had cysts both in the liver and in the lungs. Fertile cysts were found in 10 (13.2%) out of the 76 positive buffaloes. The sequencing of the mitochondrial cytochrome C oxidase subunit 1 (CO1) gene of the 48 hydatid cysts produced sequences of 419 bp for each sample analysed. For 33 samples, alignment of the obtained sequences with those present in GenBank showed a total homology with the common domestic sheep strain G1; for 15 samples, sequences obtained showed 100% homology with buffalo strain G3. The findings of the present survey represent the first epidemiological and molecular comprehensive studies on CE in water buffalo from an endemic area for E. granulosus.  相似文献   

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