Associação entre otites bacterianas e infecção pelo circovírus suíno tipo 2 (PCV2) em suínos

Neste trabalho, foi estudada a ocorrência e a bacteriologia das otites purulentas em suínos apresentando três condições clínico-patológicas distintas: síndrome multissistêmica do definhamento dos suínos (SMDS), crescimento retardado (porém sem apresentação de sinais clínicos da SMDS) e crescimento normal. No total, foram examinados 385 animais com idade de 60-130 dias, sendo a confirmação do diagnóstico da SMDS obtida pelos sinais clínicos e achados patológicos e pela detecção de antígenos virais por imunoistoquímica. De 242 animais com a SMDS, 57 (23,5%) apresentaram lesões purulentas no ouvido médio. Entre os 119 animais de baixo desenvolvimento, apenas 1 (0,7%) apresentou a lesão. Não foram detectadas lesões macroscópicas no ouvido médio dos 24 animais com crescimento normal. Os agentes isolados com maior freqüência foram Arcanobacterium pyogenes (43%), Streptococcus α-hemolíticos (37,2%) e Pasteurella multocida (27,9%) dos 86 ouvidos submetidos à bacteriologia. A alta ocorrência de lesões purulentas no ouvido médio de animais com a SMDS sugere que a infecção pelo PCV2 pode tornar o suíno mais suscetível às otites bacterianas. A ocorrência reduzida dessas lesões em suínos de baixo desenvolvimento, sem a manifestação clínica da SMDS, sugere que a otite média não representa uma causa importante de mau desempenho em leitões nas fases de crescimento e terminação. O envolvimento de A. pyogenes, Streptococcus α-hemolíticos e P. multocida na maioria das lesões apontam a importância desses organismos como agentes causais de otite média em suínos.

Création d'un modèle cellulaire des voies respiratoires du porc pour étudier les effets d'une co-infection virale au virus du syndrome reproducteur et respiratoire porcin et au circovirus porcin

Le circovirus porcin de type 2 (PCV2) est un pathogène majeur pour l’industrie porcine et est associé à une longue liste de maladies associées au circovirus porcin (MACVP). Les premières tentatives pour reproduire ces maladies ont montré que le virus doit être combiné à d’autres agents pathogènes du porc ou à différents stimulants du système immunitaire. De ces agents, le virus du syndrome reproducteur et respiratoire porcin (VSRRP) est celui qui est le plus souvent co-isolé avec le PCV dans les fermes. Une grande partie des efforts faits pour étudier les interactions entre ces deux virus ont été menés in vivo. Les interactions in vitro ont jusqu’à maintenant été peu étudiées du fait qu’il n’existe pas de modèle cellulaire permettant la réplication efficace des deux virus. L’objectif de ce projet était donc de développer un modèle cellulaire propice à la réplication des deux virus et d’étudier leur interaction en co-infection. Une lignée cellulaire provenant de la trachée d’un porcelet nouveau-né (NPTr), permissive au PCV, a été génétiquement modifiée pour exprimer la protéine CD163, un récepteur majeur du VSRRP. Ce projet a montré que cette nouvelle lignée cellulaire (NPTr-CD163) est permissive au VSRRP ainsi qu’à plusieurs génotypes de PCV (PCV1, PCV2a, PCV2b et PCV1/2a). De plus, les résultats obtenus lors d’infections mixtes suggèrent que la réplication du VSRRP et du PCV conditionne de façon génotype-dépendante celle du PCV puisque la réplication du PCV1 est inhibée en présence de VSRRP, alors que celle du PCV2b est significativement augmentée dans les mêmes conditions. Ni la mortalité cellulaire, ni la réponse cellulaire en cytokines n’a permis d’expliquer ces résultats. La modulation de la réplication du PCV par le VSRRP serait donc liée à un mécanisme spécifique qui demeure inconnu. De plus, cet effet varierait en fonction du génotype de PCV.

Effect of deoxynivalenol (DON) mycotoxin on in vivo and in vitro porcine circovirus type 2 infections.

Deoxynivalenol (DON) is a mycotoxin produced by Fusarium spp and is a common contaminant of grains in North America. Among farm animals, swine are the most susceptible to DON because it markedly reduces feed intake and decreases weight gain. Porcine circovirus type 2 (PCV2) is the main causative agent of several syndromes in weaning piglets collectively known as porcine circovirus-associated disease (PCVAD). The objectives of this study were to investigate the impact of DON on PCV2 replication in NPTr permissive cell line, and to determine eventual potentiating effects of DON on PCV2 infection in pigs. Noninfected and infected cells with PCV2 were treated with increasing concentrations of DON (0, 70, 140, 280, 560, 1200 ng/mL) and cell survival and virus titer were evaluated 72 h postinfection. Thirty commercial piglets were randomly divided into 3 experimental groups of 10 animals based on DON content of served diets (0, 2.5 and 3.5 mg/kg DON). All groups were further divided into subgroups of 6 pigs and were inoculated with PCV2b virus. The remaining pigs (control) were sham-inoculated with PBS. In vitro results showed that low concentrations of DON could potentially increase PCV2 replication depending on virus genotype. In vivo results showed that even though viremia and lung viral load tend to be higher in animal ingesting DON contaminated diet at 2.5 mg/kg, DON had no significant effect on clinical manifestation of PCVAD in PCV2b infected animals. DON has neither in vitro nor in vivo clear potentiating effects in the development of porcine circovirus infection despite slight increases in viral replication.

Development and use of a multiplex real-time quantitative polymerase chain reaction assay for detection and differentiation of Porcine circovirus-2 genotypes 2a and 2b in an epidemiological survey.

By the end of 2004, the Canadian swine population had experienced a severe 2 increase in the incidence of Porcine circovirus-associated disease (PCVAD), a problem that was 3 associated with the emergence of a new Porcine circovirus-2 genotype (PCV-2b), previously 4 unrecovered in North America. Thus it became important to develop a diagnostic tool that could 5 differentiate between the old and new circulating genotypes (PCV-2a and -2b, respectively). 6 Consequently, a multiplex real-time quantitative polymerase chain reaction (mrtqPCR) assay that 7 could sensitively and specifically identify and differentiate PCV-2 genotypes was developed. A 8 retrospective epidemiological survey that used the mrtqPCR assay was performed to determine if 9 cofactors could affect the risk of PCVAD. From 121 PCV-2–positive cases gathered for this 10 study, 4.13%, 92.56% and 3.31% were positive for PCV-2a, PCV-2b, and both genotypes, 11 respectively. In a data analysis using univariate logistic regressions, PCVAD compatible 12 (PCVAD/c) score was significantly associated with the presence of Porcine reproductive and 13 respiratory syndrome virus (PRRSV), PRRSV viral load, PCV-2 viral load, and PCV-2 14 immunohistochemistry (IHC) results. Polytomous logistic regression analysis revealed that 15 PCVAD/c score was affected by PCV-2 viral load (P = 0.0161) and IHC (P = 0.0128), but not by 16 the PRRSV variables (P > 0.9); suggesting that mrtqPCR in tissue is a reliable alternative to IHC. 17 Logistic regression analyses revealed that PCV-2 increased the odds ratio of isolating 2 major 18 swine pathogens of the respiratory tract, Actinobacillus pleuropneumoniae and Streptococcus 19 suis serotypes 1/2, 1, 2, 3, 4, and 7, which are serotypes commonly associated with clinical 20 diseases.

Preliminary study of Porcine circovirus type 2 and Torque teno sus virus coinfection frequencies in Brazilian pig herds

Torque teno sus virus (TTSuV) is emergent in swine herds. Recent studies have shown an increased frequency of TTSuV2 in Porcine circovirus type 2 (PCV2)-associated diseases (PCVAD), which are endemic in many swine-producing countries, including Brazil. Coinfection with several other viral and bacterial agents results in an increased incidence of more severe PCVAD. Given the limited information on TTSuV and PCV2 coinfection, especially in Brazilian swine herds, this study made a preliminary estimation of the occurrence of coinfection in swine herds by testing samples from different categories. Between 2008 and 2009, 111 samples of feces and 23 serum samples from 5 swine herds were tested for PCV2 and TTSuVs and the results analyzed for associations between these agents. No significant differences in coinfection frequency were observed for PCV2 1 + TTSuV1 or for PCV2 1 TTSuV2 between nursery piglets (P = 0.730), growing pigs (P = 0.331), or sows (P = 0.472). However, a significant difference was observed for PCV2 1 TTSuV1 1 TTSuV2 between nursery piglets and growing pigs (P = 0.004; Fisher's exact test). Phylogenetic studies agreed with the grouping of TTSuV1 and TTSuV2 into 2 different clades, with no distinct pattern of clustering of these isolates with the animal categories.