New PCR primers targeting the cytochrome b gene reveal diversity of Leucocytozoon lineages in an individual host.
New PCR primers targeting the cytochrome b gene reveal diversity of Leucocytozoon lineages in an individual host.
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针对细胞色素 b 基因的新 PCR 引物揭示了个体宿主中白细胞虫谱系的多样性。
DOI：
10.1007/s00436-022-07667-5
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发表时间：
2022
期刊：
影响因子：
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通讯作者：
Hikosaka K.
中科院分区：
文献类型：
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作者：
Honjo Y;Fukumoto S;Sakamoto H;Hikosaka K.
禽血孢子虫因其分布广泛、易于检测等特点，在生态学和进化方面受到广泛关注。然而，传统的基于PCR的检测方法有时可能会低估血孢子虫混合感染，这是常见的自然人群。这种低估是由于混合感染中检测谱系的PCR灵敏度的差异。因此，我们设计了新的引物来扩增常规引物不能检测到的序列，并检查我们的引物是否可用于准确检测混合感染。从北海道捕获的32只野鸟中采集血样，用常规引物检测16只，用我们的引物检测15只。对所有扩增阳性的样本进行测序，发现常规引物检测到16%（5/32）的多重感染，其中没有一个是新的谱系，而我们的引物检测到44%（14/32）的多重感染，其中10个是新的谱系。系统发育分析表明，与常规引物相比，新引物能检测到更广泛的白细胞动物谱系。结果表明，我们的引物特别适合于揭示来自多重感染的独特菌株。首次发现同一地点同一鸟群中高度可变的多重感染。我们揭示了自然界中白细胞虫谱系的多样性高于预期，这将为更好地了解野生动物中的寄生虫多样性和宿主-载体相互作用提供更多信息。
Avian haemosporidian parasites have received considerable attention in ecology and evolution as a result of their wide distribution and ease of detection. However, conventional PCR-based detection methods may sometimes underestimate haemosporidian mixed infections, which are frequent in natural populations. This underestimation is due to differences in PCR sensitivity for detection of lineages within the mixed infections. Therefore, we designed new primers to amplify sequences that were not detected by the conventional primers and examined if our primers were useful for accurate detection of mixed infections. Blood samples were collected from 32 wild birds captured in Hokkaido, and 16 of these were positive forLeucocytozoonusing the conventional primers, while 15 were positive using our primers. All positively amplified samples were sequenced, and we found that the conventional primers detected 16% (5/32) of multiple infections and none of them was a novel lineage, whereas our primers detected 44% (14/32) of multiple infections and ten of them were novel lineages. A phylogenetic analysis showed that the new primers can detect a wide range ofLeucocytozoonlineages compared with that detected by the conventional primers. The results indicate that our primers are particularly suitable for revealing unique strains from multiple infections. Highly variable multiple infections in the same population of birds at the same location were found for the first time. We revealed a higher diversity ofLeucocytozoonlineages in nature than expected, which would provide more information to better understand parasite diversity and host-vector interactions in wildlife.