Ruddell Brandon, Hassall Alan, Sahin Orhan, Plummer Paul J, Zhang Qijing, Kreuder Amanda J
Department of Veterinary Microbiology and Preventive Medicine, College of Veterinary Medicine, Iowa State University, Ames, Iowa, USA.
National Institute of Antimicrobial Resistance Research and Education (NIAMRRE), Iowa State University Research Park, Ames, Iowa, USA.
mSphere. 2025 Jan 28;10(1):e0083224. doi: 10.1128/msphere.00832-24. Epub 2025 Jan 8.
Post-transcriptional gene regulation by non-coding small RNAs (sRNAs) is critical for colonization and survival of enteric pathogens, including the zoonotic pathogen . In this study, we utilized IA3902 (a representative isolate of the sheep abortion clone) and W7 (a highly motile variant of NCTC 11168, a human gastroenteritis strain) to further investigate regulation by sRNA CjNC110. Both motility and autoagglutination ability were confirmed to be phenotypes of conserved regulation by CjNC110. However, we demonstrated that W7∆CjNC110 does not change chicken colonization levels compared to W7 wild type, directly contrasting IA3902∆CjNC110, which had decreased colonization ability. Subsequently, we determined strain-specific phenotype variation between W7∆CjNC110 and IA3902∆CjNC110 when examining intracellular L-methionine (L-met) levels controlled by the activated methyl cycle (AMC). We hypothesized that the presence of a secondary system for L-met production conferred by MetAB in W7 but not IA3902 might explain the difference in both chicken colonization and L-met availability. Insertion of within IA3902∆CjNC110 (naturally absent) restored intracellular L-met levels in IA3902∆CjNC110::metAB and overcame the colonization defect that resulted from mutagenesis of CjNC110 in IA3902. Deletion of metAB in W7∆CjNC110 (naturally present) led to a decrease in L-met in W7∆CjNC110∆metAB and a colonization defect which was otherwise masked in W7∆CjNC110. Our results indicate that regulation of the AMC leading to altered L-met availability is a conserved regulatory function of CjNC110 in and confirm that L-met generation via the AMC as activated by CjNC110 is critical for optimal host colonization.IMPORTANCEDuring this study, the regulatory action and conservation of function of CjNC110 between two different zoonotically important strains were examined. Critically, this work for the first time reveals regulation of L-methionine (L-met) production within the activated methyl cycle (AMC) by small RNA (sRNA) CjNC110 as a key factor driving optimal chicken colonization. As a growing body of evidence suggests that maintenance of L-met homeostasis appears to be critical for colonization, interventions targeting the AMC could provide a critical control point for therapeutic drug options to combat this zoonotic pathogen. Our results also indicate that even for conserved sRNAs such as CjNC110, strain-specific differences in phenotypes regulated by sRNAs may exist, independent of conserved regulatory action. Depending on the strain examined and accessory genomic content present, conserved regulatory actions might be masked, thus investigation in multiple strains may be warranted.
非编码小RNA（sRNA）介导的转录后基因调控对于肠道病原体（包括人畜共患病原体）的定殖和存活至关重要。在本研究中，我们利用IA3902（绵羊流产克隆的代表性分离株）和W7（人胃肠炎菌株NCTC 11168的高运动性变体）进一步研究sRNA CjNC110的调控作用。运动性和自凝集能力均被证实是CjNC110保守调控的表型。然而，我们发现与W7野生型相比，W7∆CjNC110的鸡定殖水平没有变化，这与IA3902∆CjNC110形成直接对比，后者的定殖能力下降。随后，我们在检测由活化甲基循环（AMC）控制的细胞内L-甲硫氨酸（L-met）水平时，确定了W7∆CjNC110和IA3902∆CjNC110之间的菌株特异性表型差异。我们推测，W7中由MetAB赋予的L-met产生的二级系统的存在（IA3902中不存在）可能解释了鸡定殖和L-met可用性的差异。在IA3902∆CjNC110（天然不存在）中插入 可恢复IA3902∆CjNC110::metAB中的细胞内L-met水平，并克服了IA3902中CjNC110诱变导致的定殖缺陷。在W7∆CjNC110（天然存在）中删除metAB导致W7∆CjNC110∆metAB中的L-met减少和定殖缺陷，而在W7∆CjNC110中该缺陷原本被掩盖。我们的结果表明，AMC的调控导致L-met可用性的改变是CjNC110在 中的保守调控功能，并证实由CjNC110激活的AMC产生L-met对于最佳宿主定殖至关重要。
重要性
在本研究中，研究了CjNC110在两种不同的人畜共患病重要 菌株之间的调控作用和功能保守性。至关重要的是，这项工作首次揭示了小RNA（sRNA）CjNC110对活化甲基循环（AMC）中L-甲硫氨酸（L-met）产生的调控是驱动 最佳鸡定殖的关键因素。越来越多的证据表明，维持L-met稳态似乎对 定殖至关重要，针对AMC的干预措施可能为对抗这种人畜共患病原体的治疗药物选择提供关键控制点。我们的结果还表明，即使对于像CjNC110这样的保守sRNA，sRNA调控的表型中也可能存在菌株特异性差异，与保守的调控作用无关。根据所检测的菌株和存在的辅助基因组内容，保守的调控作用可能会被掩盖，因此可能需要对多个菌株进行研究。
