Philpott Duncan, Näslund Joacim, Donadi Serena, Burimski Oksana, Lauringson Magnus, Pukk Lilian, Vasemägi Anti
Department of Aquatic Resources, Institute of Freshwater Research, Swedish University of Agricultural Sciences, Drottningholm, Sweden.
Chair of Aquaculture, Estonian University of Life Sciences, Tartu, Estonia.
J Fish Dis. 2025 Jun;48(6):e14095. doi: 10.1111/jfd.14095. Epub 2025 Feb 12.
Effective sample preservation is essential in large-scale population monitoring, particularly for molecular genetic analyses of pathogens, and for measuring disease symptoms in hosts. In such monitoring cases, disease symptoms can indicate poor habitat health, as they often coincide with elevated temperatures and suboptimal environmental conditions. This study examines the effect of two preservatives; 95% ethanol and 99% isopropanol on the assessment of proliferative kidney disease (PKD) in the renal tissue of young-of-the-year brown trout (Salmo trutta). Specifically, we studied the effect of preservatives on the physical measurement of a primary symptom of PKD, renal hyperplasia. Furthermore, we evaluated the effect of preservatives on the molecular detection and quantification of the causative PKD agent myxozoan parasite Tetracapsuloides bryosalmonae. Our results indicate that isopropanol-preserved samples exhibit greater renal tissue shrinkage, with the most pronounced differences observed in smaller fish when compared to ethanol-preserved samples. This difference in shrinkage is great enough to disguise symptomatic fish when observing renal hyperplasia with mixed storage mediums. However, both preservatives were found to be suitable for DNA extraction of sufficient quality for detection and quantification of the parasite using qPCR with no statistically significant differences in DNA yield or parasite load due to the type of preservative. We found that while ethanol is preferable for ease of dissection, isopropanol is a suitable alternative for PKD monitoring in wild fish, especially where access to ethanol may be limited. Understanding the difference in tissue shrinkage caused by the two preservatives can enable compensatory adjustment and maintain higher standards of data accuracy when assessing the severity of Tetracapsuloides bryosalmonae infection.
有效的样本保存对于大规模种群监测至关重要，特别是对于病原体的分子遗传分析以及宿主疾病症状的测量。在这类监测案例中，疾病症状可能表明栖息地健康状况不佳，因为它们常常与温度升高和环境条件欠佳同时出现。本研究考察了两种防腐剂；95%乙醇和99%异丙醇对当年生褐鳟（Salmo trutta）肾脏组织中增殖性肾脏病（PKD）评估的影响。具体而言，我们研究了防腐剂对PKD主要症状——肾脏增生的物理测量的影响。此外，我们评估了防腐剂对致病性PKD病原体粘孢子虫寄生虫脑粘体虫（Tetracapsuloides bryosalmonae）的分子检测和定量的影响。我们的结果表明，用异丙醇保存的样本肾脏组织收缩更为明显，与用乙醇保存的样本相比，在较小的鱼中观察到的差异最为显著。这种收缩差异大到足以在使用混合保存介质观察肾脏增生时掩盖有症状的鱼。然而，发现两种防腐剂都适合提取足够质量的DNA，以便使用qPCR检测和定量寄生虫，并且由于防腐剂类型的不同，在DNA产量或寄生虫载量方面没有统计学上的显著差异。我们发现，虽然乙醇便于解剖，但异丙醇是野生鱼类PKD监测的合适替代品，特别是在乙醇获取可能受限的情况下。了解两种防腐剂引起的组织收缩差异，可以在评估脑粘体虫感染的严重程度时进行补偿调整并维持更高的数据准确性标准。
