Institute of Pathology, Department of Pathobiology, University of Veterinary Medicine Vienna, Veterinaerplatz 1, 1210, Vienna, Austria.
Nature Research Centre, Akademijos 2, 08412, Vilnius, Lithuania.
Malar J. 2024 Mar 8;23(1):70. doi: 10.1186/s12936-024-04899-x.
Birds chronically infected with avian malaria parasites often show relapses of parasitaemia after latent stages marked by absence of parasites in the peripheral circulation. These relapses are assumed to result from the activation of dormant exo-erythrocytic stages produced during secondary (post-erythrocytic) merogony of avian Plasmodium spp. Yet, there is no morphological proof of persistent or dormant tissue stages in the avian host during latent infections. This study investigated persistence of Plasmodium relictum pSGS1 in birds with latent infections during winter, with the goal to detect presumed persisting tissue stages using a highly sensitive RNAscope® in situ hybridization technology.
Fourteen domestic canaries were infected with P. relictum pSGS1 by blood-inoculation in spring, and blood films examined during the first 4 months post infection, and during winter and spring of the following year. After parasitaemia was no longer detectable, half of the birds were dissected, and tissue samples investigated for persisting tissue stages using RNAscope ISH and histology. The remaining birds were blood-checked and dissected after re-appearance of parasitaemia, and their tissues equally examined.
Systematic examination of tissues showed no exo-erythrocytic stages in birds exhibiting latent infections by blood-film microscopy, indicating absence of dormant tissue stages in P. relictum pSGS1-infected canaries. Instead, RNAscope ISH revealed rare P. relictum blood stages in capillaries of various tissues and organs, demonstrating persistence of the parasites in the microvasculature. Birds examined after re-appearance of parasitemia showed higher numbers of P. relictum blood stages in both capillaries and larger blood vessels, indicating replication during early spring and re-appearance in the peripheral circulation.
The findings suggest that persistence of P. relictum pSGS1 during latent infection is mediated by continuous low-level erythrocytic merogony and possibly tissue sequestration of infected blood cells. Re-appearance of parasitaemia in spring seems to result from increased erythrocytic merogony, therefore representing recrudescence and not relapse in blood-inoculated canaries. Further, the study highlights strengths and limitations of the RNAscope ISH technology for the detection of rare parasite stages in tissues, providing directions for future research on persistence and tissue sequestration of avian malaria and related haemosporidian parasites.
慢性感染鸟类疟原虫的鸟类在潜伏期后经常出现寄生虫血症复发，潜伏期表现为外周循环中无寄生虫。这些复发被认为是由于在鸟类疟原虫的继发（红细胞后）裂殖体期间产生的休眠的外红细胞期的激活所致。然而，在潜伏感染期间，在禽类宿主中没有持续或休眠组织期的形态学证据。本研究在冬季研究了潜伏感染的鸟类中持久性疟原虫 relicta pSGS1 的持续性，目的是使用高度敏感的 RNAscope®原位杂交技术检测假定的持续组织期。
14 只家雀通过血液接种在春季感染 P. relicta pSGS1，并在感染后 4 个月内以及次年冬季和春季检查血片。在无法检测到寄生虫血症后，一半的鸟类进行解剖，并使用 RNAscope ISH 和组织学检查组织中持续的组织期。在寄生虫血症再次出现后，对其余鸟类进行血液检查和解剖，并对其组织进行同样检查。
通过血液涂片显微镜对表现出潜伏感染的鸟类进行组织系统检查，未发现外红细胞期，表明在 P. relicta pSGS1 感染的金丝雀中不存在休眠的组织期。相反，RNAscope ISH 显示在各种组织和器官的毛细血管中罕见的疟原虫血液期，表明寄生虫在微脉管系统中的持续性。在寄生虫血症再次出现后检查的鸟类，在毛细血管和较大的血管中都显示出更多的疟原虫血液期，表明在早春进行复制并再次出现在外周循环中。
这些发现表明，在潜伏感染期间，P. relicta pSGS1 的持续性是通过连续的低水平红细胞裂殖体和可能的受感染血细胞的组织隔离介导的。春季寄生虫血症的再次出现似乎是由于红细胞裂殖体的增加，因此代表了血液接种的金丝雀的复燃而不是复发。此外，该研究强调了 RNAscope ISH 技术在组织中检测罕见寄生虫期的优势和局限性，为研究鸟类疟疾和相关血孢子虫的持续性和组织隔离提供了方向。
