Research paper
Early inflammatory response of young rabbits attending natural resistance to calicivirus (RHDV) infection

https://doi.org/10.1016/j.vetimm.2012.09.038Get rights and content

Abstract

Young rabbits (i.e. up to 4 weeks of age) are naturally resistant to infection by rabbit haemorrhagic disease virus (RHDV), the same calicivirus that kills more than 90% of adult rabbits in 3 days or less. To characterize this fascinating model of age-related natural resistance to viral infection, we have studied the kinetics (from 6 h up to 7 days) of cytokines and of leukocyte subpopulations in the liver (the target organ for calicivirus replication) and spleen (host systemic response) of RHDV infected young rabbits. Infection was associated with early (6 h) elevation of proinflammatory cytokines (TNF-α, IL-1, IFN-α, IFN-γ, IL-6, IL-8). We found that all three major leukocyte subpopulations (macrophages, B and T lymphocytes) were increased in the liver 48 h after the RHDV inoculation. At 7 days of infection, B and T lymphocytes were still elevated in the liver of the rabbits. In the spleen, both macrophages and B lymphocytes (but not T cells) were also enhanced. At 7 days, anti-RHDV specific antibodies were present in sera of all young rabbits infected by the virus. We conclude that natural resistance of young rabbits to RHDV infection is associated with a rapid and effective inflammatory response by the liver, with few hepatocytes being infected, and also with a sustained elevation in local and systemic B and T cells.

Introduction

Rabbit haemorrhagic disease (RHD) is a widespread deadly disease of the European rabbit (Oryctolagus cuniculus) that is caused by a member of the Caliciviridae family of viruses (Ohlinger et al., 1990, Parra and Prieto, 1990, Marcato et al., 1991). RHDV infection has been responsible for the death of millions of rabbits during the last decades, both living in the wild and raised in commercial farms (Mitro and Krauss, 1993, McIntosh et al., 2007). Interestingly, young rabbits (up to 4 weeks of age) do not develop any clinical signs of RHD after RHDV inoculation (Ferreira et al., 2004). This natural resistance of young rabbits to RHDV infection was already described decade ago during the first outbreaks of this viral disease (Liu et al., 1984). Nonetheless, there is no information on the host inflammatory response that attends the acquisition of resistance to RHDV by young rabbits; this issue is the goal of the herein investigation.

We have reported before that RHDV infected young rabbits showed lymphocytic inflammatory infiltrates in the liver (Ferreira et al., 2005, Ferreira et al., 2006). We have also shown that infected young rabbits presented specific antiviral antibodies and that these antibodies, upon adoptive transfer, will protect adult rabbits from RHD (Ferreira et al., 2008).

We investigate here the early inflammatory response to RHDV by young rabbits that are naturally resistant to the virus. We document the kinetics, in liver and spleen, of leukocyte subpopulations and of serum cytokines attending experimental RHDV infection of young rabbits, up to 7 days of RHDV inoculation. It is likely that this rapid and strong inflammatory response is related with the quite low number of infected cells detected in the liver of young rabbits.

Section snippets

Rabbits

A group of 53, pathogen free, New Zealand White rabbits (O. cuniculus) was purchased from a Spanish breeder (Charles River Laboratories España, SA, Spain) and used in this study when the animals were 4 weeks old. These young rabbits and their mothers were tested for seric anti-RHDV antibodies and they were all negative. The rabbits were kept under standard conditions of housing with unrestricted access to food and water; this was done according to the European Union Directive no. 86/609/CEE.

Leukocyte kinetics in liver and spleen

Liver leukocyte number was unchanged at 24 h of RHDV infection (Fig. 1B). Liver macrophages, B and T lymphocytes, all were increased at 48 h of infection. Both B and T cells were still enhanced in the liver of rabbits at 7 days of infection. A mild decrease in the number of splenic macrophages, B and T cells was observed 24 h after infection; this early decline was followed by a significant increase in B lymphocytes and in macrophages at 48 h (Fig. 1C).

Liver and spleen CD4+ and CD8+ T lymphocytes

The CD8+ T lymphocytes were increased in the

Discussion

Young rabbits show no signs of disease after RHDV infection and will acquire long-term resistance to the virus as a result of an early contact with the microorganism (Liu et al., 1984, Ferreira et al., 2005, Ferreira et al., 2008). This age-related natural resistance of rabbits to RHDV infection is in contrast with the more than 90% mortality shown by naïve adult rabbits upon infection by the same inoculum of this calicivirus (Liu et al., 1984, Ferreira et al., 2008). Our own experience with

Acknowledgements

We are grateful to Prof. Francisco Parra (University of Oviedo) for the offer of the initial sample of RHDV suspension that was used to infect the rabbits. We thank Dr. Madalena Costa and Ms. Manuela Silva for excellent technical assistance. We are also grateful to Dr. Alexandra Correia for her kind offer of the BALB/c mice which were used for the production of anti-VP60 antibodies. This work was funded by FCT – Portugal, grant no. PTDC/CVT/66656/2006, COMPETE and FEDER. Luzia Teixeira was

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