Effect of deoxynivalenol (DON) mycotoxin on in vivo and in vitro porcine circovirus type 2 infections
Introduction
Deoxynivalenol (DON), also known as vomitoxin, is the most prevalent type B trichothecene mycotoxin worldwide (Glenn, 2007). DON contaminates cereal grains following infection in the field or during storage by Fusarium spp (Jouany, 2007). Among farm animals, swine are considered the most sensitive to DON; dietary concentrations between 2 and 5 mg/kg are frequently associated with feed refusal and concentrations over 20 mg/kg induce vomiting (Bryden, 2012). Leukocytes are among the most sensitive cells to DON effects as low concentrations of this toxin upregulate immune and inflammatory genes and high concentrations trigger cell death, typically by apoptosis, which leads to immunosuppression (Pestka et al., 2004) with a potential to decrease resistance to infectious diseases.
Previous reports in mice have shown that DON could increase reovirus replication in enteric and respiratory infection models (Li et al., 2005, Li et al., 2007). In these models, DON exacerbated viral-induced inflammation and pulmonary damage by suppressing type-1 interferon (IFN) response and elevating expression of proinflammatory cytokines (Li et al., 2007). To date, no study has reported an effect of DON on viral infections in pigs.
Porcine circovirus type 2 (PCV2) is an emerging viral disease that is of a major concern for pig health and has significant economic impacts on swine industry. PCV2 is the main causative agent of several syndromes in weaning piglets collectively known as porcine circovirus-associated disease (PCVAD) (Gillespie et al., 2009). Commonly, the clinico-pathological scope of PCVAD is characterized by severe weight loss (wasting), lymphadenopathy, jaundice, diarrhea and reproductive failure (Grau-Roma et al., 2011, Segales, 2012). PCV2 is a small, non enveloped, single stranded circular DNA virus belonging to the family Circoviridae. PCV2 can be further divided into two groups known as PCV2a and PCV2b (Olvera et al., 2007). It has been previously shown that PCV2 benefit from specific conditions induced by many others viruses such as porcine reproductive and respiratory syndrome virus (PRRSV) (Allan et al., 2000, Harms et al., 2001, Opriessnig et al., 2006b, Rovira et al., 2002), porcine torque-teno virus (Ellis et al., 2008) or porcine parvovirus (Allan et al., 1999, Hasslung et al., 2005) which can promote PCVAD. Immune stimulation with drugs such as soluble protein antigen keyhole limpet hemocyanin in incomplete Freund's adjuvant (Krakowka et al., 2001) or concanavalin A (Lefebvre et al., 2008) have been shown to enhance PCV2 replication. Since DON is an immunomodulator, the question arises if it could also enhance PCV2 replication and exacerbate the clinical signs of PCV2 infection. Therefore, the objective of this study is to evaluate the impact of DON on in vitro and in vivo PCV2 pathogenesis.
Section snippets
Cell culture
Newborn pig trachea epithelial cell line (NPTr) was kindly provided by Dr M. Ferrari (Instituto Zooprofilattico Sperimental, Brescia, Italy) (Ferrari et al., 2003). Our laboratory shows in this study that NPTr cell line is permissive to PCV2 replication (Supplementary data 1). All cell culture products were purchased from Life technologies (Burlington, ON, Canada) unless otherwise specified. NPTr was cultured in Dulbecco's modified Eagle's medium (DMEM) supplemented with 10% fetal calf serum
Cell viability
Results showed that DON concentrations of 280 ng/mL and higher were significantly detrimental to the survival of noninfected NPTr cells at 72 h (Fig. 1A). These results also demonstrate that NPTr cells untreated with DON but infected with PCV2a had a lower viability compared to noninfected cells (Fig. 1B). Effect of DON depended on virus genotype because DON had no significant effect on cell viability of PCV2a infected NPTr cells for concentration between 0 and 280 ng/mL (Fig. 1C). Conversely, it
Discussion
Animal feeds are frequently contaminated with various mycotoxins produced by secondary metabolism of diverse fungal contaminants. The food and agriculture organization (FAO) estimates that as much as 25% of the world's agricultural commodities are contaminated with mycotoxins to a certain degree (Binder et al., 2007). Among these mycotoxins, deoxynivalenol (DON) also known as vomitoxin, is the most frequent mycotoxin occurring in Canadian-grown grain (Tran et al., 2012). Mycotoxicosis is often
Acknowledgements
The authors gratefully thank the Canadian Swine Research and Development Cluster for funding this project. C. Savard and C. Provost received postdoctoral fellowships from the Canadian Swine Health Board (CSHB). C. Savard received a postdocotoral fellowship from the Fonds de recherche du Québec – Nature et technologies (FRQNT). F. Alvarez received a scholarship from Fédération des Producteurs de Porcs du Québec (FPPQ). V. Pinilla received a scholarship from CRIPA, a FRQNT network. C.A. Gagnon
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