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Detection of the bovine viral diarrhoea / mucosal disease (BVD/MD) virus in tissues from aborted ruminant foetuses using immunohistochemistry  [cached]
S.M. Njiro,C.M. Nkosi
Journal of the South African Veterinary Association , 2012, DOI: 10.4102/jsava.v80i4.213
Abstract: Various tissues from aborted ruminant foetuses were collected, fixed in formalin and embedded in paraffin wax. Sections were made and exposed to a primary monoclonal antibody against the bovine viral diarrhoea / mucosal disease (BVD/MD) virus, and subsequently to a goat anti-mouse secondary antibody conjugated to horse radish peroxidase (HRP). Diaminobenzidine (DAB) was the substrate and it released a brown pigment in the tissues on reacting with the HRP in an immunohistochemistry (IHC) procedure. Of 27 aborted foetuses, an immunoperoxidase staining reaction was observed in 1 ovine and 5 bovine foetuses. The IHC procedure located BVD/MD viral antigen in a wide variety of foetal tissues including cerebral cortical neurons, the pseudostratified columnar epithelial cells lining the bronchi, alveolar lining cells and alveolar macrophages, hepatocytes, renal tubular lining cells and the Purkinje fibres in the myocardium.
Outcome of experimental porcine circovirus type 1 infections in mid-gestational porcine foetuses
Dipongkor Saha, David J Lefebvre, Richard Ducatelle, Jan V Doorsselaere, Hans J Nauwynck
BMC Veterinary Research , 2011, DOI: 10.1186/1746-6148-7-64
Abstract: Nine foetuses from three sows were inoculated at 55 days of gestation: three with 104.3 TCID50 of the PCV1 cell culture strain ATCC-CCL33, three with 104.3 TCID50 of the PCV1 field strain 3384 and three with cell culture medium (mock-inoculated). At 21 days post-inoculation, all 6 PCV1-inoculated and all 3 mock-inoculated foetuses had a normal external appearance. Microscopic lesions characterized by severe haemorrhages were observed in the lungs of two foetuses inoculated with CCL33. High PCV1 titres (up to 104.7 TCID50/g tissue) were found in the lungs of the CCL33-inoculated foetuses. All other organs of the CCL33-inoculated foetuses and all the organs of the 3384-inoculated foetuses were negative (< 101.7 TCID50/g tissue) by virus titration. PCV1-positive cells (up to 121 cells/10 mm2 in CCL33-inoculated foetuses and up to 13 cells/10 mm2 in 3384-inoculated foetuses) were found in the heart, lungs, spleen, liver, thymus and tonsils. PCR and DNA sequencing of Rep recovered CCL33 or 3384 sequences from CCL33- or 3384-inoculated foetuses, respectively.From this study, it can be concluded that cell culture PCV1 can replicate efficiently and produce pathology in the lungs of porcine foetuses inoculated at 55 days of foetal life.Porcine circovirus type 1 (PCV1) is a small, non-enveloped circular single-stranded DNA virus of the family Circoviridae. PCV1 was originally detected as a non-cytopathic contaminant of the PK-15 cell line, ATCC-CCL33 [1]. PCV1 infections are widely distributed around the world as described before [2-4]. Seroprevalence of PCV1 at herd level varies between 10% [5] and 100% [6]. Although PCV1 DNA has been isolated from lymph nodes of a piglet in France with a wasting condition [7], it is generally accepted that PCV1 is non-pathogenic to pigs [8-13]. Experimental infections with PCV1 failed to reproduce disease in pigs [8,9,14]. The distribution of PCV1 in different pig tissues after experimental infections has been demonstrated [9]. PCV1 has bee
Co-infection by porcine circovirus type 2 and porcine parvovirus in aborted fetuses and stillborn piglets in southern Brazil
Pescador, Caroline A.;Bandarra, Paulo M.;Castro, Luiza A.;Antoniassi, Nadia A.B.;Ravazzolo, Ana Paula;Sonne, Luciana;Cruz, Cláudio E.F.;Driemeier, David;
Pesquisa Veterinária Brasileira , 2007, DOI: 10.1590/S0100-736X2007001000007
Abstract: porcine circovirus types 1 and 2 (pcv1, pcv2) and porcine parvovirus (ppv) are widespread in pig populations around the world. nevertheless, only pcv2 has been associated with different clinical syndromes, thus representing a major problem to the pig industry. the association of cases of swine abortions and stillborns with pcv1 and pcv2 and ppv was studied retrospectively (2005-2007). additional pathogens were also investigated in lesioned fetuses. the studied litters included stillborn piglets and several mummified fetuses of varied sizes. ventricular dilatation, myocardial pale areas, and mesocolic edema were the gross lesions. escherichia coli was detected as co-infecting with pcv2 the cases in which mesocolic edema was seen. microscopic lesions included non-suppurative myocarditis, myocardial necrosis and fibrosis, mineralization foci and intranuclear inclusion bodies in cardiomyocytes, and interstitial mononuclear pneumonia. samples from 7 (5.78 per cent) of 121 aborted fetuses and stillborn piglets had lesions consistent with a viral cause and showed both positive anti-pcv2 immunostaining as well as pcv2-pcr. in samples from 3 (2.47 per cent) of these 7 fetuses, co-infection with ppv was confirmed by nested-pcr. both viruses were detected in fetuses at different stages of gestation. viral antigens of pcv2 were detected by immunohistochemistry mainly in macrophages and myocytes. pcv1 individually was not detected in any of these affected fetuses, but it was associated with pcv2 and/or ppv in some of them. these findings indicate that pcv2 alone or in association with ppv should be kept in mind when investigating causes of infectious abortion in pigs in brazil.
Spread of porcine circovirus associated disease (PCVAD) in Ontario (Canada) swine herds: Part I. Exploratory spatial analysis
Zvonimir Poljak, Catherine E Dewey, Thomas Rosendal, Robert M Friendship, Beth Young, Olaf Berke
BMC Veterinary Research , 2010, DOI: 10.1186/1746-6148-6-59
Abstract: The study included 278 swine herds from a large disease-monitoring project that included porcine reproductive and respiratory syndrome (PRRS) virus-positive herds identified by the diagnostic laboratory, and PRRS virus-negative herds directly from the target population. Herds were included if they had growing pigs present on-site and available geographical coordinates for the sampling site. Furthermore, herds were defined as PCVAD-positive if a producer reported an outbreak of circovirus associated disease, or as PCVAD-negative if no outbreak was noted. Spatial trend was investigated using generalized additive models and time to PCVAD outbreak in a herd using Cox's proportional hazard model; spatial and spatio-temporal clustering was explored using K-functions; and location of most likely spatial and spatio-temporal clusters was investigated using scan statistics. Over the study period, the risk of reporting a PCVAD-positive herd tended to be higher in the eastern part of the province after adjustment for herd PRRS status (P = 0.05). This was partly confirmed for spread (Partial P < 0.01). Local spread also appeared to exist, as suggested by the tentative (P = 0.06) existence of spatio-temporal clustering of PCVAD and detection of a spatio-temporal cluster (P = 0.04).In Ontario, PCVAD has shown a general trend, spreading from east-to-west. We interpret the existence of spatio-temporal clustering as evidence of spatio-temporal aggregation of PCVAD-positive cases above expectations and, together with the existence of spatio-temporal and spatial clusters, as suggestive of apparent local spread of PCVAD. Clustering was detected at small spatial and temporal scales. Other patterns of spread could not be detected; however, survival rates in discrete Ontario zones, as well as a lack of a clear spatial pattern in the most likely spatio-temporal clusters, suggest other between-herd transmission mechanisms.Initially reported in early 1990's as post weaning multi-systemic wast
Spread of porcine circovirus associated disease (PCVAD) in Ontario (Canada) swine herds: Part II. Matched case-control study
Zvonimir Poljak, Catherine E Dewey, Thomas Rosendal, Robert M Friendship, Beth Young, Olaf Berke
BMC Veterinary Research , 2010, DOI: 10.1186/1746-6148-6-58
Abstract: A time-matched case-control study was used as a study design approach, and conditional logistic regression as the analytical method. The main exposure of interest was local spread, which was defined as an unidentified mechanism of PCVAD spread between premises located within 3 kilometers of the Euclidean distance. Various modifications of variables indicative of local spread were also evaluated. The dataset contained 278 swine herds from Ontario originally sampled either from diagnostic laboratory submissions or directly from the target population. A PCVAD case was defined on the basis of the producer's recall. Existence of apparent local spread over the entire study period was confirmed (OR = 2.26, 95% CI: 1.06, 4.83), and was further identified to be time-varying in nature - herds experiencing outbreaks in the later part of the epidemic were more likely than control herds to be exposed to neighboring herds experiencing recent PCVAD outbreaks. More importantly, the pattern of local spread was driven by concurrent occurrence of PCVAD on premises under the same ownership (OREXACTwithin ownership = 25.6, 95% CI: 3.4, +inf; OREXACToutside ownership = 1.3, 95% CI: 0.45, 3.3). Other significant factors included PRRSv status of a herd (OREXACT = 1.9, 95% CI: 1.0, 3.9), after adjusting for geographical location by including the binary effect of the easting coordinate (Easting > 600 km = 1; OREXACT = 1.8, 95% CI: 0.5, 5.6).These results preclude any conclusion regarding the existence of a mechanism of local spread through airborne transmission or indirectly through contaminated fomites or vectors, as simultaneous emergence of PCVAD could also be a result of concurrent change in contributing factors due to other mechanisms within ownerships.Porcine circovirus associated disease (PCVAD), also known as porcine circovirus disease and previously as post weaning multisystemic wasting syndrome (PMWS), emerged in the early 1990's and soon became a major animal health problem in man
Prevalence of Bovine Viral Diarrhoea Virus antibodies and antigen among the aborted cows in industrial dairy cattle herds in Mashhad area of Iran  [cached]
Hashemi Tabar, G.R.,Haghparast, A.,Naseri, Z.
Archives of Razi Institute , 2011,
Abstract: The measurement of antibody responses of animals exposed to BVDV either through a natural exposure or an immunization protocol is still a standard procedure. For BVDV, the test formats have been largely limited to ELISA which is a valuable diagnostic test to measure the level of BVDV specific antibodies as well as antigen in blood samples. In the present study, 120 blood samples were collected from the cows with the history of abortion in different period of pregnancy from different industrial dairy cattle herds of Mashhad area of Iran. Also 30 samples were collected from the cows with no history of abortion as control. The presence of antibody against BVDV from the 120 serum samples was investigated by indirect ELISA. From 120 serum samples which were collected from aborted cows, 89 samples were positive (%74.16). From these positive samples, 12(13.48%), 54 (60.68%) and 23 (25.84%) samples belong to the first, second and third trimester of pregnancy, respectively. From 89 positive samples, 12 (13.48%) samples were related to stillbirth and 8 (8.99%) samples were belongs to the mummified fetus. From 89 positive samples, 71 (79.78%) were related to cattle between 2-5 years old and 18 (20.22%) were associated to cattle more than 5 years old. In control group, 20 samples (66.66%) were antibody positive. Also the presence of BVDV antigen in serum samples was investigated by Ag-capture ELISA. From 120 serum samples, 2 samples were positive (1.67%), which belongs to the second period of pregnancy. In control group, none of the samples were antigen positive. The results of this study showed that the prevalence of BVDV infection is high among the aborted cows of Mashhad area. Although this prevalence is higher than the control group, the observed difference is not significant.
Dynamics of serum antibodies to and load of porcine circovirus type 2 (PCV2) in pigs in three finishing herds, affected or not by postweaning multisystemic wasting syndrome
Inger M Brunborg, Caroline Fossum, Bj?rn Lium, Gunilla Blomqvist, Elodie Merlot, Anne J?rgensen, Lena Eliasson-Selling, Espen Rimstad, Christine M Jonassen, Per Wallgren
Acta Veterinaria Scandinavica , 2010, DOI: 10.1186/1751-0147-52-22
Abstract: Sequentially collected blood samples from 40 pigs in each of two Swedish (A and B) and one Norwegian (C) finishing herds were analysed for serum PCV2-load and -antibodies and saliva cortisol. The two Swedish herds differed in PMWS status, despite receiving animals from the same sow pool (multi-site production). However, the PMWS-deemed herd (A) had previously also received pigs from the spot market. ResultsThe initial serum PCV2 load was similar in the two Swedish herds. In herd A, it peaked after two weeks in the finishing herd and a high number of the pigs had serum PCV2 levels above 107 per ml. The antibody titres increased continually with exception for the pigs that developed PMWS, that had initially low and then declining antibody levels. Pigs in the healthy herd B also expressed high titres of antibodies to PCV2 on arrival but remained at that level throughout the study whereas the viral load steadily decreased. No PCV2 antibodies and only low amounts of PCV2 DNA were detected in serum collected during the first five weeks in the PMWS-free herd C. Thereafter a peak in serum PCV2 load accompanied by an antibody response was recorded. PCV2 from the two Swedish herds grouped into genotype PCV2b whereas the Norwegian isolate grouped into PCV2a. Cortisol levels were lower in herd C than in herds A and B.The most obvious difference between the Swedish finishing herds and the Norwegian herd was the time of infection with PCV2 in relation to the time of allocation, as well as the genotype of PCV2. Clinical PMWS was preceded by low levels of serum antibodies and a high load of PCV2 but did not develop in all such animals. It is notable that herd A became affected by PMWS after errors in management routine, emphasising the importance of proper hygiene and general disease-preventing measures.A role of porcine circovirus type 2 (PCV2) in the etiology of postweaning multisystemic wasting syndrome (PMWS) was first observed in Canada in 1991, and described in the late 1990s
Global Gene Expression Profiling of Myeloid Immune Cell Subsets in Response to In Vitro Challenge with Porcine Circovirus 2b  [PDF]
Bettina Mavrommatis, Victoria Offord, Robert Patterson, Mick Watson, Theo Kanellos, Falko Steinbach, Sylvia Grierson, Dirk Werling
PLOS ONE , 2014, DOI: 10.1371/journal.pone.0091081
Abstract: Compelling evidence suggests that the early interaction between porcine circovirus 2 (PCV-2) and the innate immune system is the key event in the pathogenesis of Post-Weaning Multisystemic Wasting Syndrome (PMWS). Furthermore, PCV2 has been detected in bone-marrow samples, potentially enabling an easy spread and reservoir for the virus. To assess the gene-expression differences induced by an in-vitro PCV2b infection in different three different myeloid innate immune cell subsets generated from the same animal, we used the Agilent Porcine Gene Expression Microarray (V2). Alveolar macrophages (AM?s), monocyte-derived dendritic cells (MoDCs) and bone-marrow cells (BMCs) were generated from each animal, and challenged with a UK-isolate of a PCV2 genotype b-strain at a MOI of 0.5. Remarkably, analysis showed a highly distinct and cell-type dependent response to PCV2b challenge. Overall, MoDCs showed the most marked response to PCV2b challenge in vitro and revealed a key role for TNF in the interaction with PCV2b, whereas only few genes were affected in BMCs and AM?s. These observations were further supported by an enrichment of genes in the downstream NF-κB Signalling pathway as well as an up regulation of genes with pro-apoptotic functions post-challenge. PCV2b challenge increases the expression of a large number of immune-related and pro-apoptotic genes mainly in MoDC, which possibly explain the increased inflammation, granulomatous inflammation and lymphocyte depletion seen in PMWS-affected pigs.
Development of a loop-mediated isothermal amplification method to rapidly detect porcine circovirus genotypes 2a and 2b  [cached]
Qiu Xiaohuo,Li Tian,Zhang Guorui,Cao Jingjing
Virology Journal , 2012, DOI: 10.1186/1743-422x-9-318
Abstract: Background Porcine circovirus type 2 (PCV2), is nowadays associated with a number of diseases known as porcine circovirus-associated diseases (PCVAD), especially postweaning multisystemic wasting syndrome (PMWS). The epidemiological investigation of PCV2 infection was usually conducted by PCR, nested PCR, PCR-RFLP, TaqMan-based assay and nucleotide sequencing. However, there is still no rapid, sensitive and practical method for detecting PCV2 genotypes. As a novel nucleic acid amplification method, the loop-mediated isothermal amplification method (LAMP) has been used to detect a variety of pathogenic microorganisms. Results Herein, a LAMP method is developed to detect the genotypes of PCV2. The diagnostic sensitivity of LAMP is 1 copy/reaction for differentiating genotypes PCV2a and PCV2b. The reaction process was completed at 65°C for 1 hour in a water bath. Cross-reactivity assay shows that this method is specific for PCV2a and PCV2b and no reactive for PCV2c and other swine-origin viruses (i.e. CSFV, PRRSV, BVDV, TGEV and PEDV, etc). Identity between LAMP and nested PCR was 92.3% on 52 field clinical samples. Conclusions LAMP method provides a rapid, sensitive, reliable way to detect PCV2a and PCV2b, and a better means for the large scale investigation of PCV2a and PCV2b infection.
Reproduction in porcine circovirus type 2 (PCV2) seropositive gilts inseminated with PCV2b spiked semen
Sarli Giuseppe,Morandi Federico,Panarese Serena,Bacci Barbara
Acta Veterinaria Scandinavica , 2012, DOI: 10.1186/1751-0147-54-51
Abstract: Background Since 1999, field evidence of transplacental infection by porcine circovirus type 2 (PCV2) and reproductive failure has been reported in pigs. The objective of this study was to evaluate the clinical and pathological consequences of PCV2 infection in conventional PCV2-seropositive gilts by insemination with PCV2b-spiked semen. Results Six PCV2 seropositive gilts were inseminated with PCV2b-supplemented semen (infected) and three animals with semen and cell culture medium (controls). Only three out of the six infected animals were pregnant by ultrasonography on day 29 after insemination, while two out of the three controls were pregnant. One control gilt aborted on day 23 after insemination but not due to PVC2. Viraemia was demonstrated in four out of six infected and in one control gilt that became infected with PCV2a. Anti-PCV2 antibody titres showed dynamic variations in the infected group throughout the study. Among infected gilts, the animal with the lowest anti-PCV2 titre (1/100) at the beginning of the experiment and another that reached a similar low value during the experiment showed evident seroconversion over time and had also PCV2 positive foetuses. One placenta displayed mild focal necrosis of the chorionic epithelium positively stained by immunohistochemistry for PCV2 antigen. Conclusions PCV2-seropositive gilts can be infected with PCV2 after intrauterine exposure and low maternal antibody titre may increase the probability of a foetal infection.
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