【摘要】：衣原體(Chlamydia spp.)是一種嚴格胞內(nèi)寄生的原核細胞微生物,革蘭氏染色陰性,鏡下外形呈現(xiàn)球狀;其能夠在雞胚和易感的細胞系中良好地生長、繁殖,生活史分為原體和始體兩個時期。衣原體不僅在自然界廣泛存在,在美洲、歐洲、亞洲、非洲和大洋洲均有感染的報道。它的宿主也十分多樣,可感染多種禽類和哺乳動物。衣原體門包括1個衣原體綱(Chlamydiia),1個衣原體目(Chlamydiales),衣原體目下設(shè)8個科,其中衣原體科(Chlamydiaceae)是與引起人和動物患病的相關(guān)衣原體種,目前總共有12個衣原體種:沙眼衣原體(C.trachomatis)、肺炎衣原體(C.pneumonia)、流產(chǎn)衣原體(C.abortus)、鸚鵡熱衣原體(C.psittaci)、豬衣原體(C.suis)、家畜衣原體(C.pecorum).貓衣原體(C.felis)、鼠衣原體(C.muridarum)、豚鼠衣原體(C.caviae)、朱鷺衣原體(C.ibidis)、鳥衣原體(C.avium)和家禽衣原體(C.gallinacea)。鸚鵡熱衣原體一直被認為是引起禽衣原體病的唯一病原,直到近幾年C.galliacea和鳥衣原體在歐洲、中國等地區(qū)被發(fā)現(xiàn)。根據(jù)當前的調(diào)查發(fā)現(xiàn),鳥衣原體在鴿子中檢出較多,而C.gallinacea在雞、火雞中廣泛流行。目前C.gallinace 在家禽中的流行率甚至超過了鸚鵡熱衣原體,關(guān)于C.galliacea的報道主要是不同地區(qū)的流行狀況、宿主專嗜性和基因組學(xué)研究,但其傳播方式仍然未知。為了對C.gallinace 在雞群中的水平傳播方式進行探究,本研究利用禽用隔離器建立了水平傳播實驗?zāi)Ｐ?使用通氣管連接排氣口和進氣口,讓SPF雞通過糞口途徑和空氣吸入途徑分別接觸經(jīng)陽性雞群排泄物污染的食物、飲水和空氣。Ⅰ號隔離器中的SPF雞可以直接接觸人工接種的帶毒雞群及其糞便,Ⅱ號隔離器為對照組,Ⅲ號隔離器中SPF雞吸入的氣體全部來自于人工接種的帶毒雞群所在的隔離器。實驗結(jié)果表明,第15天時糞口途徑感染組中所有的SPF雞都感染了 C.gallinacea,而空氣傳播組在第5、10、15、20、25天所采集的喉拭子與泄殖腔拭子樣品檢測C.galliacea均為陰性,對照組中兩種拭子樣品檢測也均為陰性,因此證明C.galliacea能夠通過糞口途徑傳播,但不能夠通過空氣傳播。隨后,分別從人工感染組、直接接觸組和空氣傳播組中隨機選取5只雞檢測臟器中C.gallinacea分布。結(jié)果顯示空氣傳播組中所有臟器均為陰性,而人工感染組和直接接觸組中臟器C.gallinacea陽性個數(shù)如下:心臟(2/10)、肺臟(2/10)、脾臟(3/10)、腎臟(1/10)、腺胃(3/10)、十二指腸(7/10)、直腸(10/10)、泄殖腔(10/10)、卵巢(1/10),其中十二指腸、直腸、泄殖腔C.gallinacea的陽性率和平均基因拷貝數(shù)均高于其他臟器。為了對C.gallinacea在雞群中的垂直傳播方式進行研究,本研究采集了江蘇省邵伯鎮(zhèn)某種雞場100只種母雞泄殖腔拭子樣品、28只種公雞泄殖腔拭子及精液樣品;采集其所產(chǎn)種蛋的蛋殼拭子樣品294份,其中128枚種蛋(18枚為孵化9日后經(jīng)照蛋發(fā)現(xiàn)未受精蛋)收集蛋清、蛋黃樣品;55枚種蛋經(jīng)孵化得到19日齡的雞胚采集心、肝、脾、肺、腎、腸等臟器樣品,應(yīng)用基于衣原體23S rRNA基因的FRET-qPCR方法進行C.gallinacea的檢測。研究表明,在該種雞場種母雞泄殖腔拭子樣品的陽性率為75%(75/100),而公雞精液中樣品的陽性率為0%(0/28),公雞泄殖腔拭子樣品陽性率為0%(0/28)。294枚種蛋樣品中,蛋殼拭子的陽性率為97.62%(287/294),蛋清陽性率為7%(9/128),蛋黃陽性率為5.4%(7/128),并且所有蛋黃樣品檢測為陽性的種蛋,其蛋清也是陽性。所檢雞胚臟器中,心臟的陽性率為9%(5/55),肝臟為5.5%(3/55),脾臟為12.7%(7/55),肺臟為11%(6/55),腎臟為 14.5%(8/55),腸為 7.3%(4/55)。本研究是首次對C.gallinacea傳播方式進行的探索,通過建立水平傳播實驗?zāi)Ｐ?驗證了糞口途徑為Cgalinacea在雞群中水平傳播的主要方式。同時,通過檢測種雞場中種雞喉拭子、泄殖腔拭子、種蛋、雞胚臟器等樣品,提示C.gallinacea可能通過蛋殼滲透等方式在親代和子代之間發(fā)生垂直傳播。
[Abstract]:Chlamydia spp. ) It is a strictly intracellularly parasitic prokaryotic cell microorganism which is negative in gram staining and has a spherical shape under the microscopic shape, and can be well grown and propagated in the chicken embryo and the easy-to-sense cell line, and the life history is divided into a plasma and a starting body for two periods. Chlamydia is not only widely present in nature, but also in the Americas, Europe, Asia, Africa and Oceania. Its host is also very diverse and can infect a variety of birds and mammals. Chlamydia trachomatis (Chlamydia trachomatis),1 Chlamydia,1 Chlamydia, and 8 families in the order of chlamydia, of which the family of Chlamydia is the type of chlamydia associated with the cause of the disease of humans and animals, and there are currently 12 Chlamydia species: Chlamydia trachomatis, and C. pneumonia. Chlamydia trachomatis (C. abnorus), Chlamydia psittaci, C. suis, and C. pecium. Chlamydia trachomatis (C. felis), C. muridum, C. caviae, C. bialis, C. avium and C. golinacea. Chlamydia psittaci has been thought to be the only pathogen that causes the disease of the birds, and it has been found in the regions of Europe, China, and the like in the last few years. According to the current survey, the number of chlamydia is detected in pigeons, and C. golinacea is widely used in chickens and turkeys. At present, the prevalence of C. gallinace in poultry is even higher than that of Chlamydia psittaci, and the report on C. galactia is mainly the prevailing situation in different regions, the specificity of the host and the study of genomics, but the mode of transmission is still unknown. In order to explore the way of the horizontal propagation of C. gallace in the chicken group, the present study uses the isolator to establish the horizontal propagation experimental model, and uses the vent pipe to connect the exhaust port and the air inlet. And the SPF chickens respectively contact the food, the drinking water and the air which are polluted by the excrement of the positive chicken group through the way of the manure mouth and the air suction way. The SPF chickens in the No.1 isolator can be directly contacted with the artificially-inoculated chicken flocks and their feces, and the No. II isolator is the control group, and the gas sucked from the SPF chickens in the III-type isolator is all from the isolator with the artificial inoculation of the infected chicken group. The results of the experiment show that all SPF chickens in the faecal route infection group were infected with C. golinacea in the 15th day, while the throat swabs collected in the 5th, 10th, 15th, 20th and 25th days of the air-spreading group were negative, and both of the two swabs in the control group were negative. It has thus been shown that C. galliacea can be spread through the faecal pathway, but can not be propagated through air. Then,5 chickens were randomly selected from the artificial infection group, the direct contact group and the air propagation group to detect the distribution of C. golinacea in the organs. The results showed that all the organs in the air communication group were negative, and the number of the C. golinacea positive number in the artificial infection group and the direct contact group were as follows: heart (2/10), lung (2/10), spleen (3/10), kidney (1/10), glandular stomach (3/10), duodenum (7/10) and rectum (10/10). The positive rate and the average gene copy number of the cloaca (10/10) and the ovary (1/10) were higher than those of other organs. In order to study the vertical propagation of C. gallinacea in the chicken group, a sample of 100 hens from a certain chicken farm in the Shab town of Jiangsu province was collected,28 species of cock cloaca and semen were collected, and 294 samples of the egg shell swab samples from which the eggs were produced were collected. The method comprises the following steps: (1) collecting the egg white and the egg yolk sample by 128 seed eggs (18 of which are after hatching for 9 days), collecting the egg white and the egg yolk sample, and hatching to obtain the 19-day-old chicken embryo collecting heart, the liver, the spleen, the lung, the kidney, the intestine and the like, The FRET-qPCR method based on the 23S rRNA gene of Chlamydia trachomatis was used for the detection of C. golinacea. The positive rate of the sample was 75% (75/100), the positive rate of the sample in the rooster was 0% (0/28), and the positive rate of the sample in the cock was 0% (0/28). The positive rate of the egg shell swabs was 97.62% (287/294), the positive rate of egg white was 7% (9/128), and the positive rate of egg yolk was 5.4% (7/128). And all the egg yolk samples are detected as positive seed eggs, and the egg white is also positive. The positive rate of the heart was 9% (5/55), the liver was 5.5% (3/55), the spleen was 12.7% (7/55), the lung was 11% (6/55), the kidney was 14.5% (8/55) and the intestine was 7.3% (4/55). This study was the first to explore the mode of C. golinacea, and through the establishment of the experimental model of horizontal propagation, the main way of the level of Cgallinea in the chicken population was verified. At the same time, by detecting the samples of the chicken's throat swab, the cloaca swab, the seed egg, the chicken embryo and the like in the chicken farm, it is suggested that C. gallinacea can spread vertically between the parent and the offspring through the way of egg shell penetration.
【學(xué)位授予單位】：揚州大學(xué)
【學(xué)位級別】：碩士
【學(xué)位授予年份】：2017
【分類號】：S858.31

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