Dinámica de la infección por el circovirus porcino tipo 2 y títulos de anticuerpos neutralizantes en las cerdas de reemplazo subclinicamente infectadas, y el. Effects of adjuvants on porcine circovirus type 2-associated lesions. Efectos de los adyuvantes en las lesiones asociadas con el circovirus porcino tipo 2. Estudios posteriores evidenciaron abundante presencia de una variante de circovirus porcino (PCV) en lesiones de tejido linfoide (Daft y col., ; Clark,
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Porcine circovirus – Wikipedia
The diagnosis of PMWS is a difficult task and must follow three criteria: The presence of PCV2 in lymphoid tissues must be demonstrated by in situ hybridization or immunohistochemical methods. The in situ hybridization is a more complex and expense compared to other diagnostic tools, on the other hand, one problem concerning the immunohistochemical methods for PMWS diagnostic is the lack of a commercial anti-PCV2 peroxidase conjugate; therefore, the aim of this work was to obtain a polyclonal-antibody-immunoperoxidase-conjugate for the PCV2 specific detection.
An anti-PCV2-peroxidase conjugated for the PCV2 specific detection was obtained based on the use of the available commercial vaccine against PCV2 as immunogenic inoculation for producing a polyclonal antibody in rabbits.
The conjugate obtained was able to discriminate between PCV2 and PCV1 infections and a high sensitivity and specificity of the conjugate were observed. PCV2 is nowadays accepted as the essential infectious agent of postweaning multisystemic wasting syndrome PMWS 4 porcine dermatitis and nephropathy syndrome PDNS5,6and has been also associated to reproductive failure 7,8,9. All these syndrome were porcinoo grouped as porcine circovirus diseases PCVD The PMWS is the most important disease within PCVD due to this syndrome disease causes severe economic losses in consequences of an increased mortality rates and reduced feed conversion efficiency in weaning and fattening pigs at the worldwide In addition, PMWS is considered an immunosu-ppressant disease, thence facilitates the infection with opportunist pathogens affecting the swine herd health status The diagnosis of PMWS is difficult and must follow three criteria: The presence of PCV2 DNA or antigen in lymphoid tissues must be demonstrated by in situ hybridization and immunohistochemical methods 10,12, In spite pocrino that in situ hybridization has recently become increasingly important in diagnostic procedures 14,15,16the use of this technique remains restricted to a few diagnostic laboratories because of its greater technical oprcino and expense compared to other diagnostic tools.
In the case of the immunohistochemical although a certain expertise is required, this is a cost effective and easy method that could be implemented in several laboratories for the PMWS diagnostic. Nevertheless, one problem concerning the immunohistochemistry methods for the PMWS diagnostic is the lack of a commercial anti-PCV2 peroxidase conjugate, for this reason, several laboratories has developed its own anti-PCV2 conjugate for this purpose 17, Therefore, the aim of this work was to obtain a polyclonal-antibody-immunoperoxidase-conjugate for the PCV2 specific detection.
Three-millilitres of the virus propagated was added to 20 mL of this cell suspension and seeded in 10 mL volumes into two 25 cm2 cell culture flasks. The primer pair targeted an amplicon of bp of the ORF2.
The PCR reaction was done under the conditions described by Sandvik et al. Polyclonal antiserum A rabbit polyclonal antiserum was produced at the Institute, briefly: A booster inoculation was carried out 3 weeks later at same inoculum and via. In order to improve affinity of the antiserum a third inoculation at 4 weeks after a second inoculation was performed. The animals were monitored; blood samples were taken before each immunization and evaluated on PK15 monolayer infected with PCV2 porciino an immunoperoxidase monolayer assay described by Fort et al.
Rabbits were euthanized, and serum was harvested 2 weeks after the final immunization and anti-PCV2 antiserum was assessed. IgG purification and conjugated The immunoglobulin using a protein A-Sepharose Amersham, Pharmacia column chromatography were purified following standard protocols for this method The concentration of the immunoglobulin purified was calculated using the expression .
A immunoperoxidase monolayer assay as described by Fort et al. The PCV2 is considered an emerging pathogen, causing economically circovjrus diseases worldwide 25, This virus reported by Meehan et al. The fact that for the PMWS diagnosis the presence of the PCV2 within these lesions in moderate or high amounts of the virus must be demonstrated by immunohistochemical methods 10,12 as well as the lack of commercial anti-PCV2-peroxidase conjugate imply the requirement by the laboratories involved in the PMWS diagnosis to develop their own anti-PCV2-peroxidase conjugate.
The use of the killed PCV chimeric vaccine for obtaining polyclonal antibodies against PCV2 protein capside described here is a useful strategy that circovieus be assume for those laboratories with problems in cell cultures technologies or because of the complicated PCV2 isolation and multiplication, hence to obtain enough viral amount to be used as immunogenic inoculation will be a very difficult task.
The PCV2 infection and replication in cell culture occur to very low titer of the virus 29thus with the purpose of increasing the viral amount different strategies has been described 29, Nevertheless, the fact circovurus the PCV2 multiplication in cell culture is carried out to very low amount of the virus suggests that the anti-PCV2-peroxidase conjugated obtained has a high sensitivity and could be able porcinno distinguish between subclinical PCV2 infection circvoirus PMWS condition. The high sensitivity and specificity of the anti-PCV2-peroxidase conjugate obtained in this work will allow perform the PMWS diagnosis in the future.
Nevertheless a further evaluation of this conjugate in tissues samples will be needed to carry out. In summary, in this paper we described the anti-PCV2-peroxidase conjugate obtainment based on the use of the available commercial vaccine against PCV2 as immunogenic inoculation to produce a polyclonal antibody in rabbits.
The obtained conjugate was curcovirus to discriminate between PCV2 and PCV1 infections and a high sensitivity of the conjugate was observed. Seventh report of the ICTV. Academic Press, San Diego.
Hoogland MJ. Effects of adjuvants on porcine circovirus type 2-associated lesions.
Pathogenesis of porcine circovirus; experimental infections of colostrum deprived piglets and examination of pig foetal material. Studies on epidemiology and pathogenicity of porcine circovirus. Harding J, Clark E. Recognizing and diagnosing postweaning multisystemic wasting syndrome PMWS.
Circovirus porcino: un virus pequeño que genera un gran problema
Identification of porcine circovirus in tissues of pigs with porcine dermatitis and nephropathysyndrome. Myocarditis and abortion associated with intrauterine infection of sows with porcine circovirus J Vet Diagn Invest. Multiple porcine circovirus 2 associated abortion and reproductive failure in a multiple-site swine production unit.
Studies on pathogenic aspects of the post-weaning multisystemic wasting syndrome PMWS. Anim Health Res Rev. Circocirus of postweaning multisystemic wasting syndrome caused by Porcine circovirus 2: Postweaning multisystemic wasting syndrome: Post-weaning multisystemic wasting syndrome PMWS in pigs: A proposal on porcine circovirus type 2 PCV2 genotype definitions and circovirrus relation with postweaning multisystemic wasting syndrome PMWS occurrence.
Pathological, immunohistochemical, and in situ hybridization studies of natural cases of postweaning multisystemic circvoirus syndrome PMWS in pigs. Production, characterisation and applications of monoclonal antibodies to porcine circovirus 2. Development of a polyclonal-antibody-based immunohistochemical method for the detection of type 2 porcine circovirus in formalin-fixed, paraffin-embedded tissue.
First report of porcine circovirus type 2 infections in Cuba. Clrcovirus and genetic typing of porcine circovirus DNA isolated from archived paraffin embedded pig tissues.
Methods in Molecular Biology. Recent developments in the periodate method of conjugating horseradish peroxidase HRPO to antibodies. Immunofluorescence and related staining techniques. Genetic characterization of type 2 porcine circovirus PCV-2 from citcovirus with postweaning multisystemic wasting syndrome in different geographic regions of North America and development of a differential PCR-restriction fragment length polymorphism assay to detect and differentiate between infections with PCV-1 and PCV Experimental infection of 3-week-old conventional colostrum-fed pigs with porcine circovirus type 2 and porcine parvovirus.
Characterization of novel circovirus DNAs circovirue with wasting syndromes in pigs. Nucleotide sequence of porcine circovirus associated with postweaning multisystemic wasting syndrome in pigs. Isolation of porcine circovirus-like virus from pigs with a wasting disease in the USA and Europe.
Porcine circovirus 2 infection of epithelial cells is clathrin- caveolae- and dynamin-independent, actin and Rho-GTPase-mediated, and enhanced by cholesterol depletion. Recibido ; Aprobado