Bandara Aloka B, Zuo Ziwei, McCutcheon Kelly, Ramachandran Siddharth, Heflin James R, Inzana Thomas J
Department of Biomedical Sciences and Pathobiology, Virginia-Maryland College of Veterinary Medicine (Bandara, Inzana).
Department of Physics, College of Science (Zuo, McCutcheon, Heflin).
J Vet Diagn Invest. 2018 Nov;30(6):821-829. doi: 10.1177/1040638718803665. Epub 2018 Sep 28.
Histophilus somni is an opportunistic pathogen responsible for respiratory and systemic diseases of cattle and sheep. Rapid and accurate detection of H. somni is essential to distinguish H. somni from other potential pathogens for proper control and treatment of infections. Nanomaterial optical fiber biosensors (NOFS) recognize analyte interactions, such as DNA hybridization, with high specificity and sensitivity, and were applied to detect H. somni DNA in culture and clinical samples. An ionic self-assembled multilayer (ISAM) film was fabricated on a long-period grating optical fiber, and a biotinylated, nucleotide probe complementary to the H. somni 16S rDNA gene was coupled to the ISAM film. Exposure of the ISAM::probe to ⩾100 killed cells of H. somni strain 2336 without DNA amplification resulted in attenuation of light transmission of ⩾9.4%. Exposure of the complexed fiber to Escherichia coli or non- H. somni species of Pasteurellaceae reduced light transmission by ⩽3.4%. Exposure of the ISAM::probe to blood, bronchoalveolar fluid, or spleen from mice or calves infected with H. somni resulted in ⩾24.3% transmission attenuation. The assay correctly detected all 6 strains of H. somni tested from culture, or tissues from 3 separate mice and calves tested in duplicate. Six heterologous strains (representing 6 genera) reacted at below the cutoff value of 4.87% attenuation of light transmission. NOFS detected at least 100 H. somni cells without DNA amplification within 45 min with high specificity. Although different fibers could vary in signal sensitivity, this did not affect the sensitivity or specificity of the assay.
睡眠嗜血杆菌是一种机会致病菌，可引发牛和羊的呼吸道及全身性疾病。快速准确地检测睡眠嗜血杆菌对于将其与其他潜在病原体区分开来，从而对感染进行恰当的控制和治疗至关重要。纳米材料光纤生物传感器（NOFS）能够以高特异性和灵敏度识别诸如DNA杂交等分析物相互作用，并被用于检测培养物和临床样本中的睡眠嗜血杆菌DNA。在长周期光栅光纤上制备了离子自组装多层（ISAM）膜，并将与睡眠嗜血杆菌16S rDNA基因互补的生物素化核苷酸探针偶联到ISAM膜上。在不进行DNA扩增的情况下，将ISAM::探针暴露于≥100个睡眠嗜血杆菌菌株2336的死菌中，导致光传输衰减≥9.4%。将复合光纤暴露于大肠杆菌或巴斯德菌科非睡眠嗜血杆菌属的细菌中，光传输减少≤3.4%。将ISAM::探针暴露于感染了睡眠嗜血杆菌的小鼠或小牛的血液、支气管肺泡液或脾脏中，导致光传输衰减≥24.3%。该检测方法正确地检测出了从培养物中测试的所有6株睡眠嗜血杆菌，或从3只单独的小鼠和小牛的组织中进行重复测试的结果。6株异源菌株（代表6个属）的反应低于光传输衰减4.87% 的临界值。NOFS在45分钟内无需DNA扩增即可高特异性地检测到至少100个睡眠嗜血杆菌细胞。尽管不同的光纤在信号灵敏度上可能存在差异，但这并不影响检测方法的灵敏度或特异性。
