Harwood James D
Department of Entomology, University of Kentucky, S-225 Agricultural Science Center North, Lexington, KY 40546-0091, USA.
Environ Entomol. 2008 Aug;37(4):990-5. doi: 10.1603/0046-225x(2008)37[990:asnsap]2.0.co;2.
Molecular analysis of predation enables accurate and reliable elucidation of trophic linkages in complex food webs, but identifying the strength of such interactions can be subject to error. Currently two techniques dominate: monoclonal antibody-based enzyme-linked immunosorbent assay (ELISA) and polymerase chain reaction (PCR). Although the optimization and characterization of these systems ensures their sensitivity and specificity, predator collection protocols such as sweep-netting and vacuum sampling could overestimate feeding rates because of surface-level contamination, yielding positive reactivity or predation within the sampling device. Therefore, two sampling techniques (sweep-net sampling and hand collection) were compared within an alfalfa agroecosystem using a monoclonal antibody-based ELISA to test the hypothesis that cross-contamination is a source of error, i.e., significantly more predators (linyphiid spiders) would test positive for prey (Diptera) proteins. A concurrent study examining the viability of trapping predators into saline solution was also undertaken. No significant differences were found between the proportions of spiders screening positive for Diptera when collected by sweep-net versus hand collection, rejecting the hypothesis that sweep-netting predators for subsequent molecular gut content analysis overestimates predation frequency. ELISA was also capable of detecting prey proteins in predator guts from pitfall traps containing phosphate-buffered saline, indicating the suitability of this approach for the collection and analysis of epigeal predators. Although these results indicate that sweep netting and pitfall trapping into solution is appropriate in this predator-prey and ELISA analysis system, caution should be exercised with other interactions and PCR-based analysis. The likelihood for false-positive reactivity should therefore be considered on a case-by-case basis.
捕食作用的分子分析能够准确可靠地阐明复杂食物网中的营养联系，但确定此类相互作用的强度可能会出现误差。目前有两种技术占据主导地位：基于单克隆抗体的酶联免疫吸附测定（ELISA）和聚合酶链反应（PCR）。尽管这些系统的优化和特性确保了它们的灵敏度和特异性，但诸如扫网法和真空采样法等捕食者采集方案可能会因表面污染而高估捕食率，导致采样装置内出现阳性反应或捕食情况。因此，在苜蓿农业生态系统中，使用基于单克隆抗体的ELISA比较了两种采样技术（扫网采样和手工采集），以检验交叉污染是误差来源这一假设，即显著更多的捕食者（皿蛛科蜘蛛）对猎物（双翅目）蛋白质检测呈阳性。同时还开展了一项研究，考察将捕食者诱捕到盐溶液中的可行性。通过扫网法与手工采集的蜘蛛中，对双翅目检测呈阳性的比例之间未发现显著差异，这一结果否定了为后续分子肠道内容物分析而用扫网法采集捕食者会高估捕食频率的假设。ELISA还能够检测到来自装有磷酸盐缓冲盐水的陷阱中的捕食者肠道内的猎物蛋白质，表明这种方法适用于地表捕食者的采集和分析。尽管这些结果表明在这种捕食者 - 猎物和ELISA分析系统中，扫网法和陷阱诱捕法是合适的，但对于其他相互作用和基于PCR的分析应谨慎使用。因此，应根据具体情况考虑出现假阳性反应的可能性。
