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DNA VACCINE AGAINST PSEUDOTUBERCULOSIS IN MARINE FISH

Foreign code F140008007
File No. S2012-1123-C0
Posted date Oct 30, 2014
Country WIPO
International application number 2013JP074075
International publication number WO 2014038662
Date of international filing Sep 6, 2013
Date of international publication Mar 13, 2014
Priority data
  • P2012-198719 (Sep 10, 2012) JP
Title DNA VACCINE AGAINST PSEUDOTUBERCULOSIS IN MARINE FISH
Abstract Provided is a DNA vaccine for use in fish which induces protective immunity against pseudotuberculosis. As the active component, this DNA vaccine contains DNA containing a nucleotide sequence that encodes a polypeptide that is immunogenic against Photobacterium damselae subsp. piscicida, or contains an expression vector containing said DNA.
Outline of related art and contending technology BACKGROUND ART
Representative fish and shellfish aquaculture industry many aquatic organisms, a closed system aquaculture region of the viral disease and bacterial disease is, from the fact that the individual is present at a high density, the greater the influence of their infection, the culture becomes a serious problem in the industry.
Disease include nodules (or pasteurellosis) is, in the year 1963 (Chesapeake) white perch america chesapeake Gulf (rosin residue, synchronous America: Roccus americanus) for the first time as a large amount of the cause of death disease (non-patent document 1) have been reported. South-west Shikoku Japan 0 year 1968 years its occurrence in the culture of the debris found in fish, the culture of the following year in the western part of the year 1969 epidemic at almost no debris. In addition, not only for the debris black sea bream, sea bream, kijihata, ayu, such as and also occurs in a fish species, this disease have very strong infectivity in question from the threatening marine aquacultural (non-patent document 1).
The photo P. damselae subsp. piscicida belonging to genus bacteria, gram-negative, non-motile anaerobic bacilli exhibit short penetration (0.6-1.2x0.8-2.6 µm) in one family. 25-30 ° C. is appropriate for the growth of bacteria of the present invention, the optimal pH is 7.5-8. 0, 2-3% is optimal in a concentration of sodium chloride, ampicillin, oxolinic acid, insensitive to such as florfenicol. Present the symptoms of, before and after 1 mm in the spleen and kidney formation characterized by small white spots. Small white spot is arranged on the colonies of bacteria, many nodules surrounded by fibrous tissue forms. The formation of these bacteria to colonize on the withstand intracellular digestion by phagocytic cells, phagocytic cells within the capillaries and can be caused to proliferate within the interstitial tissue and cells form spheres (non-patent document 2) based on.
In regard to, the present disease have already been approved for an inactivated vaccine is, inactivated vaccines are difficult to induction of cellular immunity. However, the ability to induce cellular immunity is hardly developed vaccine (patent document 1 and patent document 2) does not.
Is for the prevention or treatment of bacterial infections, the vaccine is generally used. Inactivated vaccines (Japanese encephalitis, such as weil's disease), toxoid (tetanus, diphtheria and the like), attenuated vaccine (BCG, such as poliovirus), gene recombinant vaccine (such as a hepatitis virus B) and the like. And an inactivated vaccine and a detoxified exotoxin toxoid, to induce antibodies against these relatively safe vaccine. The recombinant vaccine is, compared to an inactivated vaccine, because it does not contain impurities, believed to be a safe vaccine than.
However, these vaccines can induce antibody production is in is, induced cell-mediated immunity has the drawback of difficult. In addition, the vaccine is an inactivated vaccine and attenuated, industrially protein antigen is obtained in a large amount is required, suitable pathogens is essential for growth. Further, attenuated vaccine that is acquired by the immune effector is, in many cases is maintained for a long period, side effects on the other hand, there is a risk that pointed out. And genetically engineered vaccine is an inactivated vaccine, the persistence of the antigen and shorter in the host, in need of adjuvant. These conventional type of vaccine from manufacture until inoculation to a subject, it is necessary to refrigerated storage, the increase in cost and a problem occurs that the decreased potency of.
Recently, vaccine research and development proceeds, the immunogenic protein by administration of a plasmid encoding DNA, one immune induction of a new vaccine (DNA vaccine) have been developed, the disadvantage of a conventional type such as vaccine described below have been improved. That is, DNA vaccine, as well as a humoral immune response, can be induced strong cell-mediated immunity, to confer protection from infection can be, further, can be highly purified, stable even under high temperature or room temperature, refrigerated storage is not essential that the long term storage is possible, by a genetic engineering procedure easy rapid improvement DNA vaccine, and shortening of the time spent by vaccine development and advantages.
Constituent proteins of rhabdoviruses (Rhabdovirus) of glycoprotein encoding gene by injection into the muscle, soleus of rainbow trout and stimulate an immune response is known (non-patent document 3). In addition, the vaccine for rainbow trout and soleus DNA (non-patent document 4) a report. However, in other fish species no report of vaccine DNA.
Scope of claims (In Japanese)請求の範囲 [請求項1]
 フォトバクテリウム・ダムセラエ亜種ピシシダ(Photobacterium damselae subsp. piscicida)に対する免疫原性ポリペプチドをコードするヌクレオチド配列を含むDNA、若しくはその配列をヒラメのコドン使用頻度を元に改変したヌクレオチド配列を含むDNA、又は前記DNAを含む発現ベクターを有効成分として含むことを特徴とする、魚類用DNAワクチン。

[請求項2]
 前記免疫原性ポリペプチドが、フォトバクテリウム・ダムセラエ亜種ピシシダのppa1、ppa2およびppars1からなる群から選んだ遺伝子にコードされているポリペプチド又はその部分断片である、請求項1に記載の魚類用DNAワクチン。

[請求項3]
 前記免疫原性ポリペプチドが、配列番号2、4又は6で表されるアミノ酸配列からなるポリペプチド又はその部分断片である、請求項2に記載の魚類用DNAワクチン。

[請求項4]
 前記免疫原性ポリペプチドが、(1)配列番号2、4又は6で表されるアミノ酸配列を含むポリペプチド、(2)配列番号2、4又は6で表されるアミノ酸配列において、1又は複数個のアミノ酸が欠失、置換、又は付加されたアミノ酸配列を含み、しかも、フォトバクテリウム・ダムセラエ亜種ピシシダに対する免疫原性を有する改変ポリペプチド、若しくは(3)配列番号2,5又は8で表されるアミノ酸配列との同一性が80%以上であり、しかも、フォトバクテリウム・ダムセラエ亜種ピスシシダに対する免疫原性を有する相同ポリペプチド、又はそれらの部分断片である、請求項1に記載の魚類用DNAワクチン。

[請求項5]
 前記ヌクレオチド配列が、(1)配列番号1、3、5、7、9又は11で表されるヌクレオチド配列、若しくは(2)1、3、5、7、9又は11で表されるヌクレオチド配列との相同性が80%以上であり、しかも、フォトバクテリウム・ダムセラエ亜種ピシシダに対する免疫原性を有するポリペプチドをコードするヌクレオチド配列、又はそれらの部分配列である、請求項1に記載の魚類用DNAワクチン。

[請求項6]
 前記発現ベクターが、天然型遺伝子を含むプラスミドwild-ppa1、wild-ppa2若しくはwild-pars1、又は改変遺伝子を含むプラスミドopt-ppa1、opt-ppa2若しくはopt-ppars1である、請求項1に記載の魚類用DNAワクチン。

[請求項7]
 請求項1~6のいずれか一項に記載の魚類用DNAワクチンを魚に投与することを特徴とする、類結節症の予防又は治療方法。

[請求項8]
 前記魚がスズキ目、フグ目又はキュウリウオ目に属する魚である、請求項7に記載の方法。

[請求項9]
 請求項1~6のいずれか一項に記載の魚類用DNAワクチンの、海産魚の類結節症に対する免疫応答の誘発への使用。

  • Applicant
  • ※All designated countries except for US in the data before July 2012
  • NATIONAL UNIVERSITY CORPORATION TOKYO UNIVERSITY OF MARINE SCIENCE AND TECHNOLOGY
  • Inventor
  • HIRONO Ikuo
  • KONDO Hidehiro
  • YAMASHITA Kozue
IPC(International Patent Classification)
Specified countries National States: AE AG AL AM AO AT AU AZ BA BB BG BH BN BR BW BY BZ CA CH CL CN CO CR CU CZ DE DK DM DO DZ EC EE EG ES FI GB GD GE GH GM GT HN HR HU ID IL IN IS JP KE KG KN KP KR KZ LA LC LK LR LS LT LU LY MA MD ME MG MK MN MW MX MY MZ NA NG NI NO NZ OM PA PE PG PH PL PT QA RO RS RU RW SA SC SD SE SG SK SL SM ST SV SY TH TJ TM TN TR TT TZ UA UG US UZ VC VN ZA ZM ZW
ARIPO: BW GH GM KE LR LS MW MZ NA RW SD SL SZ TZ UG ZM ZW
EAPO: AM AZ BY KG KZ RU TJ TM
EPO: AL AT BE BG CH CY CZ DE DK EE ES FI FR GB GR HR HU IE IS IT LT LU LV MC MK MT NL NO PL PT RO RS SE SI SK SM TR
OAPI: BF BJ CF CG CI CM GA GN GQ GW KM ML MR NE SN TD TG

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