Potomac horse fever (PHF), a severe and frequently fatal febrile diarrheal disease, has been known to be caused only by Neorickettsia risticii, an endosymbiont of digenean trematodes. Here, we report the cell culture isolation of a new Neorickettsia species found in two locations in eastern Ontario, Canada, in 2016 and 2017 (in addition to 10 variable strains of N. risticii) from N. risticii PCR-negative horses with clinical signs of PHF. Gene sequences of 16S rRNA and the major surface antigen P51 of this new Neorickettsia species were distinct from those of all previously characterized N. risticii strains and Neorickettsia species, except for those from an uncharacterized Neorickettsia species culture isolate from a horse with PHF in northern Ohio in 1991. The new Neorickettsia species nonetheless had the characteristic intramolecular repeats within strain-specific antigen 3 (Ssa3), which were found in all sequenced Ssa3s of N. risticii strains. Experimental inoculation of two naive ponies with the new Neorickettsia species produced severe and subclinical PHF, respectively, and the bacteria were reisolated from both of them, fulfilling Koch's postulates. Serological assay titers against the new Neorickettsia species were higher than those against N. risticii Whole-genome sequence analysis of the new Neorickettsia species revealed unique features of this bacterium compared with N. risticii We propose to classify this new bacterium as Neorickettsia finleia sp. nov. This finding will improve the laboratory diagnosis of and vaccine for PHF, environmental risk assessment of PHF, and understanding of PHF pathogenesis and Neorickettsia biology in general.IMPORTANCE Despite the detection of Neorickettsia species DNA sequences in various trematode species and their hosts, only three Neorickettsia species have been cell culture isolated and whole-genome sequenced and are known to infect mammals and/or cause disease. The molecular mechanisms that enable the obligatory intracellular bacterium Neorickettsia to colonize trematodes and to horizontally transmit from trematodes to mammals, as well as the virulence factors associated with specific mammalian hosts, are unknown. Potomac horse fever (PHF) is a severe and acute systemic infectious disease of horses, with clinical signs that include diarrhea. Neorickettsia risticii is the only known bacterial species that causes PHF. Ingestion of insects harboring N. risticii-infected trematodes by horses leads to PHF. Our discovery of a new Neorickettsia species that causes PHF and whole-genome sequence analysis of this bacterium will improve laboratory diagnosis and vaccine development for PHF and will contribute to our understanding of Neorickettsia ecology, pathogenesis, and biology.

中文翻译：

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导致波托马克马热的新型新立克次体物种的分离和分子分析。

波托马克马热（PHF）是一种严重的且经常致命的高热性腹泻病，仅由双歧杆菌的内生共生体新立克次氏菌引起。在这里，我们报告了2016年和2017年在加拿大东部安大略省的两个地点（除了10个变种的N.risticii菌株）发现的一种新的新立克次体物种的细胞培养物分离结果，该菌株带有临床症状的N.risticii PCR阴性马PHF。该新新立克次体物种的16S rRNA和主要表面抗原P51的基因序列与以前表征的所有N. risticii菌株和新立克次体物种的基因序列均不同，除了来自俄亥俄州北部一匹患有PHF的马的未鉴定的新立克次氏种培养物1991年。尽管如此，新的立克次体物种在菌株特异性抗原3（Ssa3）中仍具有特征性的分子内重复序列，该序列在猪链球菌菌株的所有测序Ssa3中均发现。用新的新立克次体物种对两个幼稚小马进行实验接种分别产生了严重的和亚临床的PHF，并从这两个小动物中重新分离出细菌，满足了科赫的假设。针对新新立克次氏菌的血清学滴定度高于针对新立克次氏菌的效价。对新新立克次氏菌种的全基因组序列分析揭示了该细菌与新立克次体相比的独特特征。我们建议将该新细菌分类为新立克次体finleia sp。十一月 这一发现将改善PHF的实验室诊断和疫苗，PHF的环境风险评估，重要事项尽管检测到各种吸虫物种及其宿主中的新立克次体物种DNA序列，但仅分离了三种新立克次体物种进行了细胞培养并进行了全基因组测序，并已知可感染哺乳动物和/或引起疾病。使必不可少的细胞内细菌新立克次体定殖于吸虫和从吸虫水平传播到哺乳动物的分子机制，以及与特定哺乳动物宿主有关的致病因子，尚不清楚。波托马克马热（PHF）是一种严重和急性的马全身性传染病，临床症状包括腹泻。Risticii新立克次体是唯一已知引起PHF的细菌。吞食带有N的昆虫。被马感染鼻息肉的吸虫导致PHF。我们发现了导致PHF的新立克次体物种并对该细菌进行了全基因组序列分析，这将改善PHF的实验室诊断和疫苗开发，并将有助于我们对新立克次体的生态学，发病机理和生物学认识。