School of Veterinary Medicine and Biomedical Sciences, Institute for Agriculture and Natural Resources, University of Nebraska-Lincoln, Lincoln, NE, United States.
U.S. Meat Animal Research Center, United States Department of Agriculture, Agricultural Research Service, Clay Center, NE, United States.
Front Cell Infect Microbiol. 2021 Mar 16;11:632647. doi: 10.3389/fcimb.2021.632647. eCollection 2021.
is the bacterium most often cultured from ocular lesions of cattle with infectious bovine keratoconjunctivitis, also known as bovine pinkeye. Some strains of contain operons encoding for a repeats-in-toxin (RTX) toxin, which is a known virulence factor of multiple veterinary pathogens. We explored the utility of MALDI-TOF MS and biomarker detection models to classify the presence or absence of an RTX phenotype in . Ninety strains that had undergone whole genome sequencing were classified by the presence or absence of complete RTX operons and confirmed with a visual assessment of hemolysis on blood agar. Strains were grown on Tryptic Soy Agar (TSA) with 5% sheep blood, TSA with 5% bovine blood that was supplemented with 10% fetal bovine serum, 10 mmol/LCaCl, or both. The formulations were designed to determine the influence of growth media on toxin production or activity, as calcium ions are required for toxin secretion and activity. Mass spectra were obtained for strains grown on each agar formulation and biomarker models were developed using ClinProTools 3.0 software. The most accurate model was developed using spectra from strains grown on TSA with 5% bovine blood and supplemented with CaCl, which had a sensitivity and specificity of 93.3% and 73.3%, respectively, regarding RTX phenotype classification. The same biomarker model algorithm developed from strains grown on TSA with 5% sheep blood had a substantially lower sensitivity and specificity of 68.0% and 52.0%, respectively. Our results indicate that MALDI-TOF MS biomarker models can accurately classify strains of regarding the presence or absence of RTX toxin operons and that agar media modifications improve the accuracy of these models.
是从患有传染性牛角膜结膜炎（又称牛红眼病）的牛眼部病变中最常培养出来的细菌。一些 菌株含有编码重复毒素（RTX）毒素的操纵子，RTX 毒素是多种兽医病原体的已知毒力因子。我们探讨了 MALDI-TOF MS 和生物标志物检测模型在分类 中 RTX 表型存在或不存在的应用。对 90 株进行全基因组测序的菌株，根据完整 RTX 操纵子的存在与否进行分类，并通过在血琼脂上观察溶血来确认。将菌株在添加有 5%绵羊血的胰蛋白酶大豆琼脂（TSA）、添加有 10%胎牛血清和 10mmol/L CaCl 的 5%牛血 TSA 以及同时添加两者的 TSA 上培养。这些配方旨在确定生长培养基对毒素产生或活性的影响，因为钙离子是毒素分泌和活性所必需的。为在每种琼脂配方上生长的菌株获得质谱，并使用 ClinProTools 3.0 软件开发生物标志物模型。使用在添加有 CaCl 的 5%牛血 TSA 上生长的菌株的光谱开发的最准确模型，对 RTX 表型分类的灵敏度和特异性分别为 93.3%和 73.3%。从在添加有 CaCl 的 5%绵羊血 TSA 上生长的菌株开发的相同生物标志物模型算法的灵敏度和特异性分别为 68.0%和 52.0%，要低得多。我们的结果表明，MALDI-TOF MS 生物标志物模型可以准确地对 中是否存在 RTX 毒素操纵子进行分类，并且琼脂培养基的修饰可以提高这些模型的准确性。
