The identification of sandfly species, from an area of Argentina with endemic leishmaniasis, by the PCR-based analysis of the gene coding for 18S ribosomal RNA
The identification of sandfly species, from an area of Argentina with endemic leishmaniasis, by the PCR-based analysis of the gene coding for 18S ribosomal RNA
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DOI：
10.1179/136485907x156988
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发表时间：
2007-04-01
影响因子：
--
通讯作者：
Hashiguchi, Y.
中科院分区：
文献类型：
--
作者：
Barroso, P. A.;Marco, J. D.;Hashiguchi, Y.
阿根廷萨尔塔省北部奥安省的里奥布兰科周围地区是美洲皮膜利什曼病的流行地区。为了促进该地区 Lutzomyia 物种的鉴定，通过结合 PCR 和限制性片段长度多态性 (RFLP) 分析，探索了香农陷阱捕获的白蛉 18S 核糖体 RNA (rRNA) 的编码基因序列。 PCR 产物使用了专为本研究开发的两个引物（Lu.18S 1S 和 Lu.18S AR），被克隆到商业载体（pGEM-T Easy）中，以便研究它们的核苷酸序列。在RFLP分析中，用AfaI和HapII限制性内切酶单消化和双消化的产物通过在3%或4%琼脂糖中电泳来分离。结合苍蝇形态学研究的结果，所得的 DNA 片段模式足以识别大多数捕获的白蛉为 Lu。内瓦伊。虽有另外两个物种，鲁。科特莱齐和卢。 sallesi 被收集，它们相对稀有，只能从形态上进行鉴定。用 AfaI 或 HapII 对 18S-rRNA 基因序列进行单次消化似乎足以用于 Lu 的鉴定。 neivai 来自萨尔塔省北部，以及其他几种 Lutzomyia 物种。
The area around Rio Blanco, in the OrAn department in the north of the Argentinian province of Salta, is endemic for American tegumentary leishmaniasis. In an attempt to facilitate the identification of the Lutzomyia species in this area, sequences of the gene coding for the 18S ribosomal RNA (rRNA) of sandflies caught in a Shannon trap were explored, by a combination of PCR and analysis of restriction-fragment-length polymorphism (RFLP). The products from the PCR, which employed two primers developed specifically for this study (Lu.18S 1S and Lu.18S AR), were cloned into a commercial vector (pGEM-T Easy) so that their nucleotide sequences could be investigated. In the RFLP analysis, the products of single and double digestion with the AfaI and HapII restriction enzymes were separated by electrophoresis in 3% or 4% agarose. Taken together with the results of a morphological investigation of the flies, the resultant DNA fragment patterns were sufficient to identify most of the sandflies caught as Lu. neivai. Although two other species, Lu. cortelezzii and Lu. sallesi, were collected, they were relatively rare and only identified morphologically. A single digestion of the 18S-rRNA gene sequences with AfaI or HapII appeared sufficient and useful for the identification of Lu. neivai from the north of Salta province, and for several other Lutzomyia species.