Assessment of the genetic diversity among arcobacters isolated from poultry products by using two PCR-based typing methods
Assessment of the genetic diversity among arcobacters isolated from poultry products by using two PCR-based typing methods
复制标题
DOI：
10.1128/aem.68.5.2172-2178.2002
复制
发表时间：
2002-05-01
影响因子：
4.4
通讯作者：
Vandamme, P
中科院分区：
文献类型：
--
作者：
Houf, K;De Zutter, L;Vandamme, P
在这项研究中，肠杆菌重复基因间一致性PCR（ERIC-PCR）和随机扩增多态性DNA PCR（RAPD-PCR）的特征进行了优化，弓形杆菌布氏，嗜冷弓形杆菌，斯氏弓形杆菌。此外，一个简单而快速的DNA提取方法进行了测试，用于两个分型程序。两种方法均具有良好的分型能力和区分能力，但ERIC-PCR指纹图谱比RAPD-PCR指纹图谱更复杂、重复性更好。未稀释煮沸的细胞悬液作为DNA模板的使用被认为是非常有用的ERIC-PCR。因此，优选通过ERIC-PCR方法对大量弓形杆菌属分离株进行表征。尽管DNA模板量的调整和标准化是必要的，但随后可以通过RAPD-PCR来表征产生几乎相同的ERIC指纹的分离物。在本研究的第二部分中，通过使用两种分型方法评估了肉鸡尸体上存在的弓形杆菌的基因型多样性。直接分离富集后，对24份样品的228个培养物进行了检查。采用多重PCR技术对分离株进行鉴定，结果表明，分离株为A. butzleri（n 182）和A.嗜冷菌（n = 46）。共131种（91 A. butzleri型和40个A.嗜冷菌型）的分型结果在两种分型技术之间没有不一致性。家禽分离株的分析表明，家禽产品不仅可能含有一个以上的物种，而且可能含有多个基因型。所有基因型仅限于一个家禽样品，只有三个基因型发现后，同时富集和直接分离。这些结果表明，在流行病学研究中，根据所使用的分离程序和分离株的数量，可以获得不同的结果。
In this study, enterobacterial repetitive intergenic consensus PCR (ERIC-PCR) and randomly amplified polymorphic DNA PCR (RAPD-PCR) were optimized for characterization of Arcobacter butzleri, Arcobacter cryaerophilus, and Arcobacter skirrowii. In addition, a simple and rapid DNA extraction method was tested for use in both typing procedures. Both methods had satisfactory typeability and discriminatory power, but the fingerprints generated with ERIC-PCR were more reproducible and complex than those obtained with RAPD-PCR. The use of nondiluted boiled cell suspensions as DNA templates was found to be very useful in ERIC-PCR. Characterization of large numbers of Arcobacter isolates is therefore preferably performed by the ERIC-PCR procedure. Isolates for which almost identical ERIC fingerprints are generated may subsequently be characterized by RAPD-PCR, although adjustment and standardization of the amount of the DNA template are necessary. In the second part of this study, the genotypic diversity of arcobacters present on broiler carcasses was assessed by using both typing methods. A total of 228 cultures from 24 samples were examined after direct isolation and enrichment. The isolates were identified by using a multiplex PCR as A. butzleri (n 182) and A. cryaerophilus (n = 46). A total of 131 types (91 A. butzleri types and 40 A. cryaerophilus types) were discerned without discordance between the two typing techniques. The analysis of the poultry isolates showed that poultry products may harbor not only more than one species but also multiple genotypes. All genotypes were confined to one poultry sample, and only three genotypes were found after simultaneous enrichment and direct isolation. These results demonstrate that different outcomes can be obtained in epidemiological studies depending on the isolation procedure used and the number of isolates characterized.