Based on MLST analyses the global population of C. difficile is distributed in eight clades, of which Clade 2 includes the “hypervirulent” NAP1/RT027/ST01 strain along with various unexplored sequence types (STs). To clarify whether this clinically relevant phenotype is a widespread feature of C. difficile Clade 2, we used the murine ileal loop model to compare the in vivo pro-inflammatory (TNF-α, IL-1β, IL-6) and oxidative stress activities (MPO) of five Clade 2 clinical C. difficile isolates from sequence types (STs) 01, 41, 67, and 252. Besides, we infected Golden Syrian hamsters with spores from these strains to determine their lethality, and obtain a histological evaluation of tissue damage, WBC counts, and serum injury biomarkers (LDH, ALT, AST, albumin, BUN, creatinine, Na+, and Cl−). Genomic distances were calculated using Mash and FastANI to explore whether the responses were dictated by phylogeny. The ST01 isolate tested ranked first in all assays, as it induced the highest overall levels of pro-inflammatory cytokines, MPO activity, epithelial damage, biochemical markers, and mortality measured in both animal models. Statistically indistinguishable or rather similar outputs were obtained for a ST67 isolate in tests such as tissue damage, neutrophils count, and lethal activity. The results recorded for the two ST41 isolates tested were of intermediate magnitude and the ST252 isolate displayed the lowest pathogenic potential in all animal experiments. This ordering matched the genomic distance of the ST01 isolate to the non-ST01 isolates. Despite their close phylogenic relatedness, our results demonstrate differences in pathogenicity and virulence levels in Clade 2 C. difficile strains, confirm the high severity of infections caused by the NAP1/RT027/ST01 strain, and highlight the importance of C. difficile typing.

中文翻译：

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体内动物模型证实了艰难梭菌 MLST 进化枝 2 中 NAP1/RT027/ST01 基因型的毒力潜力和致病性增加

基于 MLST 分析，全球艰难梭菌种群分布在八个进化枝中，其中进化枝 2 包括“高毒性”NAP1/RT027/ST01 菌株以及各种未探索的序列类型 (ST)。为了阐明这种临床相关表型是否是艰难梭菌进化枝 2 的普遍特征，我们使用鼠回肠环模型来比较体内促炎（TNF-α、IL-1β、IL-6）和氧化应激活性(MPO) 从序列类型 (ST) 01、41、67 和 252 中分离出五个 Clade 2 临床艰难梭菌分离株。此外，我们用这些菌株的孢子感染金叙利亚仓鼠以确定它们的致死率，并获得组织学评估组织损伤、WBC 计数和血清损伤生物标志物（LDH、ALT、AST、白蛋白、BUN、肌酐、Na+ 和 Cl-）。使用 Mash 和 FastANI 计算基因组距离，以探索反应是否由系统发育决定。测试的 ST01 分离株在所有测定中均排名第一，因为它在两种动物模型中均诱导了最高水平的促炎细胞因子、MPO 活性、上皮损伤、生化标志物和死亡率。在组织损伤、中性粒细胞计数和致死活性等测试中，ST67 分离株获得了统计上无法区分或相当相似的输出。测试的两种 ST41 分离株的记录结果为中等量级，而 ST252 分离株在所有动物实验中显示出最低的致病潜力。这种排序匹配了 ST01 分离株与非 ST01 分离株的基因组距离。尽管它们有着密切的系统发育相关性，