BACKGROUND Passerine birds are frequently infected with diverse haemosporidian parasites. While infections are traditionally considered benign in wild birds, recent studies demonstrated mortalities of passerine species due to exo-erythrocytic development of the parasites, which can damage organs in affected hosts. However, exo-erythrocytic development remains insufficiently investigated for most haemosporidian species and thus little is known about the virulence of tissue stages in wild passerine birds. The aim of the present study was to investigate natural haemosporidian infections in deceased Eurasian blackbirds (Turdus merula) and song thrushes (Turdus philomelos) and to determine parasite burden and associated histological effects. METHODS For molecular analysis, blood and tissue samples from 306 thrushes were screened for Plasmodium, Haemoproteus and Leucocytozoon parasites by nested PCR. For the detection of parasite stages in organ samples, tissue sections were subjected to chromogenic in situ hybridization (CISH) using genus- and species-specific probes targeting the rRNAs of parasites. Exo-erythrocytic parasite burden was semi-quantitatively assessed and histological lesions were evaluated in haematoxylin-eosin-stained sections. RESULTS By PCR, 179 of 277 Eurasian blackbirds and 15 of 29 song thrushes were positive for haemosporidians. Parasites of all three genera were detected, with Plasmodium matutinum LINN1 and Plasmodium vaughani SYAT05 showing the highest prevalence. CISH revealed significant differences in exo-erythrocytic parasite burden between lineages in Eurasian blackbirds, with P. matutinum LINN1 frequently causing high exo-erythrocytic parasite burdens in various organs that were associated with histological alterations. Song thrushes infected with P. matutinum LINN1 and birds infected with other haemosporidian lineages showed mostly low exo-erythrocytic parasite burdens. Two Eurasian blackbirds infected with Leucocytozoon sp. TUMER01 showed megalomeronts in various organs that were associated with inflammatory reactions and necroses. CONCLUSION This study suggests that P. matutinum LINN1, a common lineage among native thrushes, regularly causes high exo-erythrocytic parasite burdens in Eurasian blackbirds, which may result in disease and mortalities, indicating its high pathogenic potential. The findings further illustrate that the same parasite lineage may show different levels of virulence in related bird species which should be considered when assessing the pathogenicity of haemosporidian parasite species. Finally, the study provides evidence of virulent Leucocytozoon sp. TUMER01 infections in two Eurasian blackbirds caused by megalomeront formation.

中文翻译：

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野生欧亚黑鸟（Turdus merula）和鹅口疮（T. philomelos）中的血孢子虫：一项原位杂交研究，重点是外红细胞寄生虫的负担。

背景技术雀形目鸟类经常感染多种血孢寄生虫。尽管传统上将感染视为野生鸟类的良性疾病，但最近的研究表明，由于寄生虫的外红细胞发育，会导致雀形目物种死亡，这可能会损害受影响宿主的器官。但是，对于大多数血孢菌属物种，外红细胞的发育仍未得到充分研究，因此对野生雀形目鸟类组织阶段的毒性了解甚少。本研究的目的是调查已故的欧亚黑鸟（Turdus merula）和鹅口疮（Turdus philomelos）中的自然血友病感染，并确定寄生虫负担和相关的组织学影响。方法为了进行分子分析，从306个鹅口疮的血液和组织样本中筛选了疟原虫，巢式PCR检测变形杆菌和白血球菌的寄生虫。为了检测器官样品中的寄生虫阶段，使用针对寄生虫rRNA的属和种特异性探针对组织切片进行显色原位杂交（CISH）。在苏木精-伊红染色的切片中半定量评估外红细胞寄生虫负担，并评估组织学病变。结果通过PCR检测，在277个欧亚黑鸟中有179个在29个鹅口疮中有15个为血孢子虫阳性。检测到所有三个属的寄生虫，其中疟原虫LINN1和沃氏疟原虫SYAT05显示最高的流行率。CISH揭示了欧亚黑鸟与P之间系外红细胞寄生虫负担的显着差异。matutinum LINN1经常在与组织学改变相关的各个器官中引起高的外红细胞寄生虫负担。感染了P. matutinum LINN1的鹅口疮鹅和感染了其他血吸虫谱系的鸟类显示出较低的外红细胞寄生虫负担。两只被Leucocytozoon sp。感染的欧亚黑鸟。TUMER01在与炎症反应和坏死有关的各个器官中均显示出黑素细胞。结论这项研究表明，斑节对虾LINN1是天然鹅口疮中的常见血统，通常会在欧亚黑鸟中引起高的外红细胞寄生虫负担，这可能导致疾病和死亡，表明其高致病性。这些发现进一步说明，相同的寄生物谱系在相关禽类中可能显示出不同的毒力水平，在评估血吸虫病寄生虫的致病性时应考虑到这一点。最后，该研究提供了有毒的白细胞增生菌sp的证据。梅加洛美隆的形成引起了两只欧亚黑鸟的TUMER01感染。