本文選題：衣原體 + 牦牛　； 參考：《甘肅農(nóng)業(yè)大學(xué)》2017年博士論文

【摘要】：衣原體是嚴(yán)格胞內(nèi)寄生的革蘭氏陰性細菌,具有廣泛的致病性,能引起動物和人的多種疾病。感染牦牛的衣原體主要有三種,即流產(chǎn)衣原體(C.abortus)、鸚鵡熱衣原體(C.psittaci)、以及反芻動物衣原體(C.pecorum),其中流產(chǎn)衣原體感染可引起牦牛流產(chǎn)和生殖障礙性疾病。衣原體病導(dǎo)致牦牛的流產(chǎn)嚴(yán)重阻礙了牦牛相關(guān)產(chǎn)業(yè)的發(fā)展,給牧區(qū)人民帶來了巨大的經(jīng)濟損失。因此,對于牦牛衣原體病的防控具有重要的公共衛(wèi)生及經(jīng)濟意義。本研究組織開展了牦牛衣原體病流行病學(xué)調(diào)查,采集了青海省多個縣的牦牛血清和組織樣品,通過間接血凝試驗(IHA)和PCR檢測,結(jié)果顯示:2014年間接血凝試驗陽性率為24.02%,PCR檢測陽性率為23.8%;2015年間接血凝試驗陽性率為55.76%,PCR檢測陽性率為41.67%。表明青藏高原相關(guān)區(qū)域牦牛感染衣原體的形勢非常嚴(yán)峻。本研究對牦牛流產(chǎn)組織樣本進行了分離鑒定,采用雞胚卵黃囊接種培養(yǎng)法從貴南縣(GN)和海晏縣(HY)共分離到6株衣原體,經(jīng)PCR-RFLP鑒定,GN-1、GN-4、GN-6以及HY-1與流產(chǎn)衣原體標(biāo)準(zhǔn)株SX5的特征一致,為流產(chǎn)衣原體。同時,有兩株流產(chǎn)衣原體毒株連續(xù)傳代至第18代,繁殖滴度為10-10,且穩(wěn)定。本研究以分離株GN-6作為制苗菌株,進行了牦牛流產(chǎn)衣原體滅活苗的研究,以BALB/c小鼠為實驗動物進行免疫試驗。以接種雞胚卵黃囊培養(yǎng)分離得到的牦牛流產(chǎn)衣原體作為抗原,經(jīng)甲醛滅活,以206為佐劑制成牦牛流產(chǎn)衣原體滅活疫苗。經(jīng)過小鼠免疫試驗證明:免疫小鼠的抗體亞型測定結(jié)果為以κ鏈輕鏈為主IgG1型、血清與淋巴細胞培養(yǎng)上清中的細胞因子IFN-γ含量明顯升高、DTH檢測結(jié)果顯示足墊明顯增厚、淋巴細胞增殖系數(shù)升高。結(jié)果表明免疫小鼠體內(nèi)產(chǎn)生了Th1型細胞免疫應(yīng)答,對衣原體感染后的有效清除起到一定作用�？贵w中和試驗結(jié)果顯示,免疫小鼠血清中形成了能夠中和衣原體的保護性抗體。攻毒試驗結(jié)果表明,免疫組小鼠能夠有效抵抗衣原體的感染。以上結(jié)果表明小鼠在接種了該疫苗以后,誘導(dǎo)機體產(chǎn)生特異性細胞免疫應(yīng)答和體液免疫應(yīng)答。本研究對研制的牦牛流產(chǎn)衣原體疫苗進行了本動物試驗。牦牛流產(chǎn)衣原體滅活疫苗安全試驗結(jié)果表明這種衣原體滅活疫苗安全可靠;通過最小免疫劑量試驗和免疫效力檢測證明,免疫劑量為牦牛犢1ml、成年牦牛2ml,即可達到良好的免疫效果。疫苗的免疫持續(xù)期檢測結(jié)果顯示,本研究所研制的牦牛流產(chǎn)衣原體疫苗免疫后210天時平均免疫抗體滴度保持在1:256以上。此外,疫苗的保存期試驗?zāi)壳罢谶M行中,將疫苗在4℃條件下保存一年,已于2016年8月進行免疫,免疫后抗體效價較好,達到了預(yù)期的免疫效果。本研究在青藏高原地區(qū)開展了牦牛衣原體的流行病學(xué)調(diào)查,成功分離鑒定到4株流產(chǎn)衣原體,制備了具有良好免疫效果的流產(chǎn)衣原體疫苗,為牦牛衣原體病的防控奠定了基礎(chǔ)。
[Abstract]:Chlamydia chlamydia is a strictly cytosolic gram-negative bacteria with a wide range of pathogenicity and can cause a variety of animal and human diseases. There are mainly three kinds of chlamydia infection in yak, I. E. C. abortus, C. psittacius, and C. pecorumus, among which the infection of chlamydia miscarriage can cause abortion and reproductive disorders in yak. The abortion of yak caused by chlamydia has seriously hindered the development of yak related industries and brought huge economic losses to the people in pastoral areas. Therefore, the prevention and control of Chlamydia yak chlamydia has important public health and economic significance. In this study, the epidemiological investigation of yak chlamydia was carried out, and yak serum and tissue samples were collected from several counties of Qinghai Province, and detected by indirect hemagglutination test (IHA) and PCR. The results showed that the positive rate of indirect hemagglutination test in 2014 was 24.02% and that of indirect hemagglutination test in 2015 was 55.76% and 41.67% respectively. The results showed that the situation of Yak infected with Chlamydia was very serious in Qinghai-Tibet Plateau. In this study, six strains of Chlamydia chlamydia were isolated from chicken embryo yolk sac inoculation and culture from Guinan County (GNN) and Haiyan County (HYY). The characteristics of GN-1GN-4GN-6 and HY-1 were consistent with that of Chlamydia abortus standard strain SX5. For chlamydia miscarriage. At the same time, two strains of chlamydia abortus were subcultured to the 18th generation, the reproductive titer was 10-10 and stable. In this study, the inactivated Chlamydia abortus vaccine of yak was studied with GN-6 as the vaccine preparation strain, and the BALB/c mice were used as experimental animals to carry out the immune test. Using Chlamydia abortus isolated from yak embryo yolk sac culture as antigen, Chlamydia abortus inactivated vaccine was prepared by formaldehyde inactivation and 206 as adjuvant. The antibody subtype of immunized mice was mainly 魏 chain light chain IgG1, and the level of cytokine IFN- 緯 in the supernatant of serum and lymphocyte culture was significantly increased. Lymphocyte proliferation coefficient increased. The results showed that the immune response of Th1 type cells was produced in the immunized mice, which played a certain role in the effective clearance of chlamydia infection. Antibody neutralization test showed that protective antibodies against chlamydia were formed in serum of immunized mice. The results showed that the immunized mice could resist chlamydia infection effectively. The above results suggest that the specific cellular and humoral immune responses were induced after the mice were inoculated with the vaccine. In this study, the Yak Chlamydia abortus vaccine was tested. The safety test of the inactivated Chlamydia chlamydia vaccine in yak showed that the vaccine was safe and reliable. The immune dose was 1 ml for yak calf and 2 ml for adult yak. The results of immunization duration test showed that the average antibody titer of Chlamydia abortus vaccine was above 1: 256 at 210 days after immunization. In addition, the preservation period of the vaccine is currently under way. The vaccine was immunized at 4 鈩，

本文編號：1907013