Soft Tissue Digestion of Paradiplozoon vaalense for SEM of Sclerites and Simultaneous Molecular Analysis
Soft Tissue Digestion of Paradiplozoon vaalense for SEM of Sclerites and Simultaneous Molecular Analysis
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DOI：
10.1645/14-521.1
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发表时间：
2015-02-01
影响因子：
1.3
通讯作者：
Avenant-Oldewage, A.
中科院分区：
文献类型：
--
作者：
Dos Santos, Q. M.;Avenant-Oldewage, A.
大多数单殖动物的分类主要基于大小、形状和附着片的排列。当使用光学显微镜研究时，这些结构经常被模糊或误解，导致关于定义字符的混淆。扫描电子显微镜（SEM）主要用于研究较小的单殖吸虫中的附着骨片，重点是钩和锚。在梁龙科中，扫描电镜还没有被用来研究haptoral片。利用新的和改进的技术，在南非收集的二倍体骨片成功地研究了扫描电镜。使用来自DNA提取试剂盒的消化缓冲液消化周围组织，并使用聚赖氨酸包被的载玻片和无菌载玻片来限制骨片的移动和损失，后者更加用户友好。除了使用SEM成功地可视化骨片之外，来自少至一半的接合器的消化组织为分子表征提供了可行的遗传物质。从这里提出的结果，使用扫描电镜，包括那些轴承夹，研究较大的单殖吸虫的骨片是一个可行的可能性，为今后的研究。此外，这种方法可能是有益的研究其他，非haptoral骨片，如在其他家庭的monogeneans的cirri。在这项研究中，野Labeo capensis被认为是一个有效的主机的Paradiplozoon vaalense在一个地区的瓦尔河的模式主机，野Labeo umbratus，似乎是缺席。
Classification of most monogeneans is primarily based on size, shape, and arrangement of haptoral sclerites. These structures are often obscured or misinterpreted when studied using light microscopy, leading to confusion regarding defining characters. Scanning electron microscopy (SEM) has predominantly been used to study haptoral sclerites in smaller monogeneans, focusing on hooks and anchors. In the Diplozoidae, SEM has not been used to study haptoral sclerites. Using new and modified techniques, the sclerites of diplozoids collected in South Africa were successfully studied using SEM. The digestion buffer from a DNA extraction kit was used to digest the surrounding tissue, and Poly-Llysine-coated and concavity slides were employed to limit the movement and loss of sclerites, with the latter being more user-friendly. In addition to the success of visualizing the sclerites using SEM, the digested tissue from as little as half of the haptor provided viable genetic material for molecular characterization. From the results presented here, the study of the sclerites of larger monogeneans using SEM, including those bearing clamps, is a viable possibility for future research. Also, this method may be beneficial for the study of other, non-haptoral sclerites, such as cirri in other families of monogeneans. During this study, Labeo capensis was noted as a valid host of Paradiplozoon vaalense in a region of the Vaal River where the type host, Labeo umbratus, appears to be absent.