鸡主要胚胎性疫病病原人工共感染的试验研究
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摘要
禽胚胎性疾病是一类由多病原引起的综合性疾病,可造成禽胚发育受阻,胚胎死亡,孵化率降低等现象,导致养禽业的重大经济损失。近几年,家禽发生多种病原共感染的现象非常普遍,多种病原共感染常常引起非典型的病变。细菌性胚胎性疫病病原种类较多,且多数是条件性致病菌,由不同的细菌多重感染诱发的疾病越来越常见。过去报道的细菌性的家禽胚胎病病因主要有鸡白痢、大肠杆菌病、霉形体病、鸡败血支原体病(Mycoplasma gallisepricum, MG)、滑液囊支原体病等。刘红芹等2006年曾报道山东某大型AA父母代肉种鸡所产种蛋中鸡白痢、鸡败血支原体、滑液囊支原体、鸡大肠杆菌和禽波氏杆菌雏率为4.50%～17.60%、死胎率为6.73%～29.40%。鸡白痢、鸡败血支原体、滑液囊支原体、鸡大肠杆菌和禽波氏杆菌(Bordetella Avium)平均阳性率分别为13.59%、12.69%、8.10%、19.62%、37.14%。经病原分离鉴定为的多重感染。孙晴等2007年也曾报道类似的情况。由此可见鸡毒支原体、鸡沙门氏菌、鸡波氏杆菌为几种常见鸡胚胎细菌性疫病的病原,因此,本研究针对这三种常见鸡胚胎细菌性疫病的病原进行了人工共感染与检测。主要对这三种病原微生物的共感染的致病性,以及间接免疫荧光(IFA)与PCR检测方法等方面进行了系统的研究,目的在于通过本研究探讨这几种病原引发疾病时造成的危害,以及发病机理,为防治和诊断禽类胚胎细菌性疫病的多重感染提供依据。
本研究分为二部分:
一鸡主要胚胎性疫病病原人工共感染致病性的研究
本研究利用三种常见并且危害严重的鸡胚胎细菌性疫病的病原,即鸡毒支原体、鸡沙门氏菌、禽波氏杆菌进行人工共感染试验,分别对11日龄SPF鸡胚(10枚/组)和1日龄SPF雏鸡(10只/组)进行三种、两种、一种病原微生物的人工感染,雏鸡每6h观察一次,鸡胚每8h观察一次,主要对生长情况,死亡情况进行统计;并对死亡雏鸡、鸡胚进行剖检,进行病原学与病理学观察。结果三种病原的共感染致病性最强,出现死亡的时间,鸡胚为8h、雏鸡为6h,死亡率也最高,雏鸡为60%,鸡胚为100%。双重感染中,鸡毒支原体与禽波氏杆菌共感染组和鸡沙门氏菌与禽波氏杆菌共感染组出现死亡时间(12h)和死亡率(50%)相同,鸡毒支原体与鸡沙门氏菌共感染组相比较,致病性较轻,但是鸡群生长发育缓慢。无论通过肉眼观察和还是病理组织学观察,三种病原共感染的雏鸡和鸡胚病变最严重,各组织器官都出现明显病变,两种病原共感染次之。总之,多种病原发生混合感染会引起更严重的病情以及更高的死亡率,并且常常引起非典型病变,难于作出诊断。
二鸡主要胚胎性疫病病原人工共感染检测方法的研究
近几年,家禽发生多种病原共感染的现象非常普遍,多种病原共感染常常引起非典型的病变,因此,发生多种病原共感染时很难通过临床症状进行诊断。为了在发生共感染后能进行快速准确的诊断,本研究通过对三种病原(鸡毒支原体、鸡沙门氏菌、禽波氏杆菌)的共感染分析研究,其目的在于研究病原的分布状况及其病原的消长规律,建立三种病原共感染的IFA、PCR等快速检测方法。结果表明,鸡毒支原体的感染气囊感染最严重、肾脏次之、其后是肝脏;禽波氏杆菌与鸡沙门氏菌在肝脏和法氏囊感染最严重、肾脏和肺脏次之;另外,对于禽波氏杆菌感染鸡,皮下也含有大量病原菌。雏鸡在感染24h内各病原含量呈快速增长趋势,36h后达到高峰,然后缓慢下降,一直持续到8周左右。在三种病原共感染的雏鸡组织中,每种病原的含量与单独感染、双重感染的含量相差不大。鸡胚在感染24h内各病原含量呈增长趋势,24h达到最高峰,24h之后缓慢下降,多种病原共感染的鸡胚病原的含量略低于相同病原单独感染的鸡胚。建立起的三种病原共感染的IFA、PCR快速检测方法特异性强、灵敏度高、结果出现快。本研究为三种病原共感染的快速诊断提供了重要的试验数据和理论依据。
The disease of avian embryo is a synthetical disease which can cause the increase of embryo death and decrease of incubation rate and is fatal to economy. In most condition, it is very common occurrence that the avians coinfect with two or more pathogens. The manner in which multiple pathogens interact is not always straightforward, however the disease of avian embryo is a synthetical disease which can cause the increase of embryo death and decrease of incubation rate and is fatal to economy. There are many species of the disease of avian embryo, and most is conditional pathogenic bacteria. Mycoplasma gallisepricum, Salmonella Avium and Bordetella Avium are three disease of avian embryo. Liu Hong-qin reported that a kind of disease appeared in one large-scale AA broiler breeder farm in Shandong province with the symptom of embryo death, increase of weak chicken, ineffective immunity, stunted growth of 30 days chicken since June, 2004. Antibody in serum was tested to the broiler breeder, 1d chicken and the commercial chicken, it was showed that the average positive rate of bordetella avium, salmonella avium, E.coli, mycoplasma were 13.59%、12.69%、8.10%、19.62% and 37.14%, respectively. Sun Qing reported that this kind of disease appeared on three parental generation branch farms in 2007. It was showed that the co-infection of bordetella avium, salmonella avium and mycoplasma was the major cause of the disease. These pathogens spread vertically from the eggs to the embryos. Thus, the study and detection aims directly at the co-infection of these three diseases of avian embryo. The pathogenic study of synthetic co-infection of bordetella avium, salmonella avium and mycoplasma was had done. And the study of the establishment of detection methods for the co-infection with IFA and PCR was had done.The research subject aims at imitation the similar problems that usually emergent in production. The discussion on the harmfulness of pathogens interact and the pathogenesis, supplying the theoretical foundation to the prevention of avian embryo in breeder farm.
The research divides two parts:
Part One: The pathogenic study of synthetic co-infection of three major bacterium pathogens of avian embryo
In an initial experiment, the effect of coinfection of Mycoplasma gallisepricum, Salmonella Avium and Bordetella Avium was examined in seventy , 1-day-old chicken (10 chicken /group) and seventy, 11-day-old embryos (10 mbryos/group) that were infected with the three pathogens or two pathogens or only one pathogens . An additional 10 chicken and 10 embryo served as sham infected controls. Observe the growth and death of the chicken (once every 6 hours) and embryo (once every 8 hours). When they die, dissect the dead bodies. Make the tissue paraffin section with the fresh tissue. The results: The thesis aims at observing the appearance and the tissue paraffin section of the co-infection.The symptom of the coinfection of the three pathogens is the most serious, the death time is the most early (the chicken is 6h, the embryo is 8h), the death rate is highest (the chicken is60%, the embryo is 100%), and the next in order is the dually infected chicken and embryo.The grous which were co-infected by MG, Bordetella avium and Bordetella avium, Salmonella have the same death time (12h) and death rate (50).
Thus, the avians coinfect with two or more pathogens can cause the increase of embryo death and decrease of incubation rate and is fatal to breeding industry. In most condition, it is a synthetical disease.It usually causes atypical pathological changes.
Part Two: The study of detection methods for synthetic co-infection of three major bacterium pathogens of avian embryo
The recent years, it is very common occurrence that the avians coinfect with two or more pathogens. In this case, It usually causes atypical pathological changes.Thus, it is difficult to do clinical diagnosis when the co-infection is emergent. In order to investigate the potential trend in disease severity after co-infection, a second experiment was performed to examine the pathogens by IFA and PCR in tissues from single and dually infection and triplicate infection. The results: the content of MG is highist in aerocyst, Salmonella Avium and Bordetella Avium is in Liver and bursa. When it is 36h, the content of pathogens is highest in chicken and ahout 24h in emobryo. Then it will reduce slowly.
The establishment of detection technique of labelled for the co-infection with IFA and PCR are simple, rapid and strong specificity.The exeperiment provides essential essay datas for collecting samples and controlling technology for the co-infection of three bacterium pathogens of avian embryo.
本研究分为二部分:
一鸡主要胚胎性疫病病原人工共感染致病性的研究
本研究利用三种常见并且危害严重的鸡胚胎细菌性疫病的病原,即鸡毒支原体、鸡沙门氏菌、禽波氏杆菌进行人工共感染试验,分别对11日龄SPF鸡胚(10枚/组)和1日龄SPF雏鸡(10只/组)进行三种、两种、一种病原微生物的人工感染,雏鸡每6h观察一次,鸡胚每8h观察一次,主要对生长情况,死亡情况进行统计;并对死亡雏鸡、鸡胚进行剖检,进行病原学与病理学观察。结果三种病原的共感染致病性最强,出现死亡的时间,鸡胚为8h、雏鸡为6h,死亡率也最高,雏鸡为60%,鸡胚为100%。双重感染中,鸡毒支原体与禽波氏杆菌共感染组和鸡沙门氏菌与禽波氏杆菌共感染组出现死亡时间(12h)和死亡率(50%)相同,鸡毒支原体与鸡沙门氏菌共感染组相比较,致病性较轻,但是鸡群生长发育缓慢。无论通过肉眼观察和还是病理组织学观察,三种病原共感染的雏鸡和鸡胚病变最严重,各组织器官都出现明显病变,两种病原共感染次之。总之,多种病原发生混合感染会引起更严重的病情以及更高的死亡率,并且常常引起非典型病变,难于作出诊断。
二鸡主要胚胎性疫病病原人工共感染检测方法的研究
近几年,家禽发生多种病原共感染的现象非常普遍,多种病原共感染常常引起非典型的病变,因此,发生多种病原共感染时很难通过临床症状进行诊断。为了在发生共感染后能进行快速准确的诊断,本研究通过对三种病原(鸡毒支原体、鸡沙门氏菌、禽波氏杆菌)的共感染分析研究,其目的在于研究病原的分布状况及其病原的消长规律,建立三种病原共感染的IFA、PCR等快速检测方法。结果表明,鸡毒支原体的感染气囊感染最严重、肾脏次之、其后是肝脏;禽波氏杆菌与鸡沙门氏菌在肝脏和法氏囊感染最严重、肾脏和肺脏次之;另外,对于禽波氏杆菌感染鸡,皮下也含有大量病原菌。雏鸡在感染24h内各病原含量呈快速增长趋势,36h后达到高峰,然后缓慢下降,一直持续到8周左右。在三种病原共感染的雏鸡组织中,每种病原的含量与单独感染、双重感染的含量相差不大。鸡胚在感染24h内各病原含量呈增长趋势,24h达到最高峰,24h之后缓慢下降,多种病原共感染的鸡胚病原的含量略低于相同病原单独感染的鸡胚。建立起的三种病原共感染的IFA、PCR快速检测方法特异性强、灵敏度高、结果出现快。本研究为三种病原共感染的快速诊断提供了重要的试验数据和理论依据。
The disease of avian embryo is a synthetical disease which can cause the increase of embryo death and decrease of incubation rate and is fatal to economy. In most condition, it is very common occurrence that the avians coinfect with two or more pathogens. The manner in which multiple pathogens interact is not always straightforward, however the disease of avian embryo is a synthetical disease which can cause the increase of embryo death and decrease of incubation rate and is fatal to economy. There are many species of the disease of avian embryo, and most is conditional pathogenic bacteria. Mycoplasma gallisepricum, Salmonella Avium and Bordetella Avium are three disease of avian embryo. Liu Hong-qin reported that a kind of disease appeared in one large-scale AA broiler breeder farm in Shandong province with the symptom of embryo death, increase of weak chicken, ineffective immunity, stunted growth of 30 days chicken since June, 2004. Antibody in serum was tested to the broiler breeder, 1d chicken and the commercial chicken, it was showed that the average positive rate of bordetella avium, salmonella avium, E.coli, mycoplasma were 13.59%、12.69%、8.10%、19.62% and 37.14%, respectively. Sun Qing reported that this kind of disease appeared on three parental generation branch farms in 2007. It was showed that the co-infection of bordetella avium, salmonella avium and mycoplasma was the major cause of the disease. These pathogens spread vertically from the eggs to the embryos. Thus, the study and detection aims directly at the co-infection of these three diseases of avian embryo. The pathogenic study of synthetic co-infection of bordetella avium, salmonella avium and mycoplasma was had done. And the study of the establishment of detection methods for the co-infection with IFA and PCR was had done.The research subject aims at imitation the similar problems that usually emergent in production. The discussion on the harmfulness of pathogens interact and the pathogenesis, supplying the theoretical foundation to the prevention of avian embryo in breeder farm.
The research divides two parts:
Part One: The pathogenic study of synthetic co-infection of three major bacterium pathogens of avian embryo
In an initial experiment, the effect of coinfection of Mycoplasma gallisepricum, Salmonella Avium and Bordetella Avium was examined in seventy , 1-day-old chicken (10 chicken /group) and seventy, 11-day-old embryos (10 mbryos/group) that were infected with the three pathogens or two pathogens or only one pathogens . An additional 10 chicken and 10 embryo served as sham infected controls. Observe the growth and death of the chicken (once every 6 hours) and embryo (once every 8 hours). When they die, dissect the dead bodies. Make the tissue paraffin section with the fresh tissue. The results: The thesis aims at observing the appearance and the tissue paraffin section of the co-infection.The symptom of the coinfection of the three pathogens is the most serious, the death time is the most early (the chicken is 6h, the embryo is 8h), the death rate is highest (the chicken is60%, the embryo is 100%), and the next in order is the dually infected chicken and embryo.The grous which were co-infected by MG, Bordetella avium and Bordetella avium, Salmonella have the same death time (12h) and death rate (50).
Thus, the avians coinfect with two or more pathogens can cause the increase of embryo death and decrease of incubation rate and is fatal to breeding industry. In most condition, it is a synthetical disease.It usually causes atypical pathological changes.
Part Two: The study of detection methods for synthetic co-infection of three major bacterium pathogens of avian embryo
The recent years, it is very common occurrence that the avians coinfect with two or more pathogens. In this case, It usually causes atypical pathological changes.Thus, it is difficult to do clinical diagnosis when the co-infection is emergent. In order to investigate the potential trend in disease severity after co-infection, a second experiment was performed to examine the pathogens by IFA and PCR in tissues from single and dually infection and triplicate infection. The results: the content of MG is highist in aerocyst, Salmonella Avium and Bordetella Avium is in Liver and bursa. When it is 36h, the content of pathogens is highest in chicken and ahout 24h in emobryo. Then it will reduce slowly.
The establishment of detection technique of labelled for the co-infection with IFA and PCR are simple, rapid and strong specificity.The exeperiment provides essential essay datas for collecting samples and controlling technology for the co-infection of three bacterium pathogens of avian embryo.
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