Please use this identifier to cite or link to this item: http://hdl.handle.net/11455/66286
標題: 利用抗體分析及RT-PCR技術建立野外豬瘟病毒感染豬隻篩檢之模式
Detedcion of the pigs infected by Classical Swine Feve(or Hog Cholera) virus of fild strain using antibody analysis and Revers Transcriptase-PCR method
作者: 陳金蘭
關鍵字: 豬瘟病毒;ELISA KIT;抗體;PCR技術
出版社: 獸醫微生物學研究所
摘要: 
一般豬32頭,SPF豬8頭;5隻一般豬及2隻SPF豬於六週齡進行一次免
疫及另5隻一般豬及2隻SPF豬於六、九週齡分別進行兔化豬瘟疫苗免疫共
兩次,並於十一週齡同時分組進行肌肉接種及同居方式感染豬瘟病毒ALD
株,另外免疫對照組10隻亦分為一次及二次免疫,有兩隻一般豬則為攻毒
對照組。所有仔豬在免疫前先行採血,測其移行抗體,免疫後定期採血,
以間接螢光染色法測定中和抗體及以兩種不同國外製造的ELISA試劑套組
檢測抗體,比較免疫與攻毒後抗體的差異性。
分別在六週齡及六、九週齡免疫兔化豬瘟疫苗,六週齡免疫注射時所
測得其移行抗體力價平均為5.31倍及2.04倍,免疫注射三週後即九週齡時
抗體無上升現象反而漸趨下降至0.87和1.89倍,經補強注射後至13週齡才
上升至32倍,上升速度緩慢,一直到上市前體重才達到最高,另免疫一次
組則持續上升到六月齡上市前體重;在試驗組中以肌肉注射攻毒組,於六
週齡免疫注射前其移行抗體力價,平均為6.95(Log 2)和3.24 (Log 2),
免疫注射三週後即九週齡時抗體無上升反呈漸趨下降現象,在病毒攻擊前
一週(第十週)才有些微的上升,攻擊後抗體產生,力價急遽向上升高,一
直維持到六月齡上市前體重。同居感染組,於六週齡免疫前其移行抗體力
價平均為10(Log 2)和4.38(Log 2),亦在攻毒後才向上升高,之後一直持
續到上市前體重。SPF豬在免疫前的移行抗體已下降至最低 (≦3 X),所
以其在免疫後中和抗體力價隨即向上升且在攻擊後保有高保護力價抗體,
一直維持到上市前的體重。未免疫對照組的兩隻仔豬,含有高移行抗體,
因為沒有免疫注射所以其移行抗體一直下降,在病毒攻擊後一週內呈現典
型豬瘟症狀,分別在9到11天內死亡。在各組實驗豬中,由荷蘭製(H)的KIT
均可測得抗體反應,在免疫前所測得的平均值均為< 30 %範圍內,經一次
免疫和二次免疫的抗體做比較其差異性並不明顯,且攻擊病毒後亦無明顯
上升,SPF豬所測的值均為負值且一次免疫和二次免疫的抗體並無差異。
僅未免疫組為> 60 %,呈陽性反應,其抗體逐漸下降。另一組瑞士製ELISA
KIT(W製),免疫前所測得平均值,在一般豬呈陽性反應,SPF豬測得平均
值不一致,且在一次與二次免疫所測的抗體平均值差異亦不顯著。
RT-PCR為實驗室診斷豬瘟方法之一,檢測血液及唾液帶病毒情形,在
檢測經攻毒後的唾液第四天即可由各實驗組中測得病毒,除了肌肉注射於
第五天才檢測到;攻毒後豬隻血液的病毒血症,攻毒前未檢測出病毒,攻
毒後,每週均可檢測到病毒,未免疫注射組,在第一週即測出有病毒血症

Fourty piglets, including 32 conventional and 8 SPF piglets,
were divided into three groups. The first group was consisted
of 10 conventional piglets. Five of them were vaccinated with
one dose of LPC vaccine at 6 weeks old and the other five piglets
were vaccinated two times at 6 and 9 weeks old. The second group,
consisted of 10 conventional and 4 SPF piglets, were divided into
two subgroups. Each subgroup contained 5 conventional and 2 SPF
piglets. One subgroup was vaccinated at 6 weeks old, and the other
subgroup was vaccinated two times at 6 and 9 weeks old.
After vaccination, all this second group piglets were
intramuscularly inoculated with the virulent ALD hog cholera virus
at 11 weeks old. The third group was consisted of 12 conventional
and 4 SPF piglets, in which 5 conventional and 2 SPF piglets were
vaccinated at 6 weeks old and the other 5 conventional and 2 SPF
piglets were vaccinated two times at 6 and 9 weeks old. Two
conventional leftovers were unvaccinated, challenged at 11 weeks
old, and housed with other vaccinated piglets of the third group
for contact infection. Serum samples from vaccinated and unvaccinated
animals were titrated with fluorescent antibody.
The geometric means of serum neutralization (SN) titer from piglets
vaccinated at 6 or 6 and 9 weeks old were 5.31X and 2.04X at 6 weeks old
(prior to vaccination), and decreased to 0.87X and 1.89X at 9 weeks old
(3 weeks post vaccination), respectively. Following the boost at 9 weeks
old, the SN titer moved to 32X at 13 weeks old and reached peaks at the
marketing time. However, the SN titers from one time vaccination animals
reached peaks at 6 months old. The geometric means of SN titers from animals
with one or two times vaccination following ALD challenge were 6.95 X and
3.24 X at 6 weeks old and decreased to basal levels at 9 weeks old, but it
increased slightly at 10 weeks old and boosted by challenge, then maintained
in high levels through the marketing time. For vaccinated and contact
infection piglets, the geometric means of SN titers were 3.10 (log 2)
and 4.38 (log 2) prior to contact infection, and it increased rapidly,
then maintained in high levels through the marketing time. In SPF piglets,
the SN titers were undetectable prior to vaccination, and boosted by vaccination
and maintained in high levels through the marketing time . The SN titers from two
unvaccinated piglets were undetectable and the animals died at 9th and 11th days
following ALD challenge.
Antibody titers test by ELISA with the (H) brand ELISA kits showed that the antibody inhibition values of each test group were below 30 prior to vaccination. There were no significant differences between one and two times vaccination. Antibody titers test with (W) brand ELISA kits indicated that the conventional piglets were positive, while SPF piglets were flatted. In additional, there were no significant differences between one and two time vaccination.
Saliva and blood samples were collected from the ALD challenge animals and performed by RT-PCR to test the carrier status. The saliva samples, collected from each challenge group, were positive in RT-PCR at the fourth day post challenge. However, the saliva samples from vaccinated animals maintained undetectable through the test time.
URI: http://hdl.handle.net/11455/66286
Appears in Collections:微生物暨公共衛生學研究所

Show full item record
 

Google ScholarTM

Check


Items in DSpace are protected by copyright, with all rights reserved, unless otherwise indicated.