Mycobacterial 70 kD heat-shock protein is an effective subunit vaccine against bovine paratuberculosis

doi:10.1016/j.vaccine.2005.12.019

Vaccine

Volume 24, Issue 14, 24 March 2006, Pages 2550-2559

https://doi.org/10.1016/j.vaccine.2005.12.019 Get rights and content

Abstract

Paratuberculosis is a chronic granulomatous inflammation of the small intestine of cattle and other ruminants, caused by infection with Mycobacterium avium ssp. paratuberculosis (MAP). The disease can be found in ruminant herds worldwide, causing substantial economic losses at farm level due to premature culling and production losses.

In previous studies, it has been shown that immune responses to recombinant MAP Hsp70 proteins were predominantly cell mediated. As protective immunity to the intracellular mycobacterial pathogens is thought to be cell-mediated in origin, we have studied the use of a recombinant MAP Hsp70 as a subunit vaccine in cattle experimentally infected with MAP.

The results of the current study demonstrate that recombinant MAP Hsp70 can be successfully used as a subunit vaccine against bovine paratuberculosis, significantly reducing shedding of bacteria in feces during the first 2 years following experimental infection.

Introduction

Paratuberculosis is a chronic granulomatous inflammation of the small intestine of cattle and other ruminants, caused by infection with Mycobacterium avium ssp. paratuberculosis (MAP). The disease can be found in ruminant herds worldwide, causing substantial economic losses at farm level due to premature culling and production losses. In addition, it may represent a potential food-safety issue by transmission of MAP, the causative organism, to humans via dairy and other products [1], [2].

Transmission of paratuberculosis involves the presence of the bacterium in several substrates. From infected animals, bacteria are mainly excreted in feces and when the disease progresses towards the clinical stage of infection also milk. Following excretion, the organism is able to survive in the environment for prolonged periods of time in substrates such as surface water and soil [1], [3].

Eradication of paratuberculosis is hampered by a lack of reliable diagnostic tools with high specificity and sensitivity and could be aided substantially with a vaccine [4]. The currently available vaccine consists of various variations of whole bacterins with adjuvants. These vaccines have been shown to have a variable efficacy in field studies [5], [6]. From these studies it has become apparent that the current vaccine prevents the occurrence of the clinical stage of the disease to a high degree, thereby limiting a substantial amount of the direct economical damage. In cattle, however, the vaccine does not prevent infection and limits the frequency of subclinically infected animals, which shed bacteria in their feces intermittently, only marginally at best. Other major drawbacks of the whole bacterin vaccines are the interference tuberculosis and paratuberculosis diagnostics and eradication due to the fact that vaccinated animals become false positive in regular tuberculosis and paratuberculosis diagnostic assays [5], [6], [7], [8], [9]. A third drawback is that the vaccine in its current formulation causes substantial local tissue reaction, in terms of prolonged swelling and granuloma formation at the site of injection [10].

Relative little effort has been put in studying recombinant protein antigens of MAP in general and in vaccine studies in particular. With previous work we have documented immune responses to mycobacterial heat-shock proteins (Hsp) in various stages of bovine paratuberculosis. The Hsp are a family of evolutionary conserved proteins, expressed in both prokaryotic and eukaryotic organisms. The expression of Hsp is upregulated during the cellular stress that occurs in both host and pathogen in response to infection and inflammation. We have previously shown that the Hsp are immunodominant antigens in bovine paratuberculosis [11], [12], similar to other mycobacterial diseases such as tuberculosis and leprosy, eliciting strong cell-mediated and antibody responses. A number of studies on mycobacterial diseases, mainly in murine model systems, have indicated a potential use of Hsp as vaccine components. Our studies indicated that immune responses to recombinant MAP Hsp70 proteins were predominantly cell mediated contrary to recombinant MAP Hsp65, which apparently induced mainly antibody responses and minor cell-mediated responses [11], [13].

As protective immunity to the intracellular mycobacterial pathogens is thought to be cell mediated in origin, we have studied the use of a recombinant MAP Hsp70 as a subunit vaccine in cattle experimentally infected with MAP. New candidate vaccines against bovine paratuberculosis should have the potential to limit transmission of infection between susceptible animals to aid in eradication strategies. In the current study we therefore studied immune parameters as well as fecal excretion in the first 2 years of experimental paratuberculosis to determine the potential of the recombinant MAP Hsp70 as a subunit vaccine in a relatively short timeframe considering the natural course bovine paratuberculosis.

Section snippets

Animals and experimental design

A total of 40 female calves (aged 29 ± 9 days at the start of the experiment) were used in the current study. The calves were raised using conventional procedures and feeds and were checked daily for general health. The calves were randomly assigned to one of the four experimental groups, according to a 2 × 2 factorial design (Table 1). Calves in groups 1 and 2 were physically separated from calves in groups 3 and 4, and rigorous hygienic measures were taken to prevent infection of the control

Observations on general health status

The growth of the animals was monitored throughout the experiment and the results indicated that there were no differences in growth between the different treatment groups (data not shown), Around day 56, an outbreak of ringworm skin disease, caused by Trichophyton verrucosum, affected all animals in groups 3 and 4; this infection resolved spontaneously. Around day 70, 16 out of 20 animals in groups 1 and 2 showed signs of a respiratory infection; while 10 cases resolved spontaneous, 6 were

Discussion

To our knowledge this is the first report describing a heat-shock protein-based subunit vaccine approach to bovine paratuberculosis, which leads to significantly decreased shedding of bacteria into the feces. Hsp have been shown to be dominant antigens for the immune system in many infectious diseases (reviewed in [21]). The Hsp70 of M. tuberculosis and M. bovis has been used previously in DNA vaccination studies, both in calves and in murine model systems, with variable success as therapeutic

Acknowledgements

The authors express their thanks to Hans Vernooij for expert help with the statistical analysis. This study was supported by a grant from the Technology Foundation (STW) of the Dutch Research Council (NWO), grant number STW-UDG5589.

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Mycobacterial 70 kD heat-shock protein is an effective subunit vaccine against bovine paratuberculosis

Abstract

Introduction

Section snippets

Animals and experimental design

Observations on general health status

Discussion

Acknowledgements

Vet Clin North Am Food Anim Pract

Vet Clin North Am Food Anim Pract

Vet Microbiol

J Dairy Sci

Immunol Today

Vet Immunol Immunopathol

Vet Microbiol

Curr Opin Immunol

J Biol Chem

Vet Microbiol

Dev Comp Immunol

Vet Immunol Immunopathol

Vet Immunol Immunopathol

Vet Immunol Immunopathol

Vet Immunol Immunopathol

Survival and dormancy of Mycobacterium avium subsp. paratuberculosis in the environment

Appl Environ Microbiol

Mycobacterium avium subsp. paratuberculosis: pathogen, pathogenesis and diagnosis

Rev Sci Tech

Immune reactions in cattle after immunization with a Mycobacterium paratuberculosis vaccine and implications for the diagnosis of M. paratuberculosis and M. bovis infections

J Vet Med B Infect Dis Vet Public Health

Use of long-term vaccination with a killed vaccine to prevent fecal shedding of Mycobacterium avium subsp. paratuberculosis in dairy herds

Am J Vet Res

Accidental self-inoculation with Mycobacterium paratuberculosis bacterin (Johne's bacterin) by veterinarians in Wisconsin

J Am Vet Med Assoc

Differential changes in heat shock protein-, lipoarabinomannan-, and purified protein derivative-specific immunoglobulin G1 and G2 isotype responses during bovine Mycobacterium avium subsp. paratuberculosis infection

Infect Immun

Progressive bovine paratuberculosis is associated with local loss of CD4(+) T cells, increased frequency of gamma delta T cells, and related changes in T-cell function

Infect Immun