Prevalence of Met-203 type spaA variant in Erysipelothrix rhusiopathiae isolates and the efficacy of swine erysipelas vaccines in Japan
Introduction
Erysipelothrix rhusiopathiae is a Gram-positive bacillus that causes erysipelas in a variety of mammals and birds, and dermatological erysipeloid disease in humans [1], [2]. Erysipelas is commonly associated with swine, and can be divided into three types by its clinical signs: acute (septicemia), sub-acute (urticaria), and chronic (arthritis, lymphadenitis, and endocarditis). Acute swine erysipelas is characterized by generalized septicemia often resulting in sudden death, and is specified as a notifiable infectious disease in Japan. Pigs infected with erysipelas are prohibited from being slaughtered or are condemned at the slaughterhouse by the Slaughterhouse Act, to prevent human consumption. The pig industry therefore suffers considerable economic loss from this disease.
For the prevention of swine erysipelas, attenuated live vaccine prepared from the acriflavine-fast attenuated Koganei strain 65–0.15 of E. rhusiopathiae has been used in Japan for many years because of its low-cost and convenience. Appropriate inoculation with the live vaccine produces a strong immunity [3], [4]. An inactivated vaccine was approved in 1997 because the live vaccine had a higher incidence of adverse events in pigs kept in a clean environment, such as specific-pathogen-free (SPF) pigs [5], [6]. Several kinds of inactivated vaccines have been approved in Japan for swine erysipelas, and their use has been increasing in recent years.
The surface protective antigen (Spa) protein of E. rhusiopathiae has been shown to be highly immunogenic and is a promising vaccine candidate for erysipelas [7], [8]. The Spa proteins of E. rhusiopathiae can be classified into 3 molecular species, named SpaA (including serotypes 1a, 1b, 2, 5, 8, 9, 12, 15, 16, 17 and N), SpaB (serotypes 4, 6, 11, 19 and 21) and SpaC (serotype 18) [8]. A mouse cross-protection study showed that three recombinant Spa proteins (rSpaA, rSpaB and rSpaC) elicited complete protection against challenge with homologous strains but that the level of protection against heterologous strains varied depending on the rSpa protein used for immunization [8]. This indicated that the Spa proteins, especially the N-terminal half of the hyper-variable region, are important for specific immunity. A recent report by To et al. [9] demonstrated that a variant isolate with aspartic acid (Asp) and methionine (Met) at the amino acid positions 195 (Asp-195) and 203 (Met-203), respectively, in the hyper-variable region of the spaA gene might be widespread among pig populations in Japan. The Met-203 isolates, therefore, differ in nucleotide sequence, in the 432-bp hypervariable region of the spaA gene, from the live vaccine strain Koganei 65-0.15 [9].
Unfortunately, according to national statistics, the morbidity rate of swine erysipelas in Japan has been increasing since 2009 [10]. The aim of this study was to characterize recently isolated E. rhusiopathiae strains, assess the prevalence and pathogenicity of the spaA variants, and re-evaluate the validity of swine erysipelas vaccines against a spaA variant field isolate.
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Bacterial strains and growth conditions
A total of 34 E. rhusiopathiae isolates were collected for this study. They were isolated from pigs with erysipelas by the Livestock Hygiene Centers from 12 prefectures (Aomori, Niigata, Yamanashi, Ibaraki, Gunma, Nagano, Mie, Gifu, Saitama, Kyoto, Fukuoka and Kumamoto) from 2008 to 2010. Twenty-one isolates originated from acute cases (septicemia), three from sub-acute (urticarial), and 10 from chronic (arthritis and endocarditis) cases (Table 1). All of the cases were identified as
Sequence of spaA genes
The sequences of the 432-bp hypervariable region of the spaA gene of the 34 field isolates were compared with that of the E. rhusiopathiae Fujisawa strain (Table 2). Based on the sequence analysis of this fragment, the 34 field isolates could be divided into three groups as follows: (i) 19 isolates with Met-203 and isoleucine at position 257 (Ile-257) classified as Group 1, (ii) 12 isolates with Ile-257 classified as Group 2, and (iii) three isolates with Ala-195 and Ile-257 classified as Group
Discussion
In Japan, serious outbreaks of swine erysipelas were noted in 1939 and 1967. In the 1967 outbreak, the prevalence rate was 0.26%, indicating that 1 in 380 pigs in Japan were infected with swine erysipelas [6]. The incidence of acute and sub-acute swine erysipelas decreased sharply after the 1970s, with the popularization of the live vaccine, but sporadic cases involving 1500–2000 pigs annually continued, even after 1996 [6], [10]. The morbidity rate of swine erysipelas, reflected in the
Acknowledgment
We would like to thank the staff of the Livestock Hygiene Service Centers for supplying the isolates of E. rhusiopathiae.
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