식품 내 병원성 대장균 O157:H7의 검출을 위한 압타머 기반의 이중기능링커 개발 및 금 나노입자 응집 기반 비색 어세이 개발

학위논문(석사) -- 서울대학교대학원 : 농업생명과학대학 농생명공학부, 2023. 2. 최영진.In this study, we developed a colorimetric assay with bi-functional gold nanoparticle (AuNP) based on aptamer for sensitive detection of Escherichia coli O157:H7 using the aggregation of AuNP linked with a complementary DNA aptamer specific to E. coli O157:H7 aptamer. AuNP was used as a colorimetric linker and probe, respectively, and E. coli O157:H7 aptamer and its complementary DNA sequences were modified on each AuNP surface. AuNP labeled with E. coli O157:H7 aptamer was used as a bi-functional linker (BL) in a colorimetric competitive assay. Colorimetric assay with BL based on aptamer detected target bacteria by sequential competitive binding between colorimetric probes, which are AuNP having complementary sequences of E. coli O157:H7 (AA, Aggregation Assistant) and E. coli O157:H7 aptamer. In addition, under optimized conditions by main testing parameters, the assay exhibited a detection range of 101 to 106 CFU/400 µL and a detection limit of 57 ± 0.5 CFU/400 µL. In addition, the developed colorimetric assay with BL based on aptamer was excellently evaluated as an on/off detection system in coconut water, a real matrix. Thus, the proposed colorimetric competitive assay with BL based on aptamer can be a proper on-site detection method of foodborne pathogens. In addition, this study provides the basis for research into developing different colorimetric assay with aptamer for pathogen analysis in real matrices. This colorimetric assay with BL based on aptamer functions by driving the system between two competitive interaction dynamics: A) affinity between the aptamer sequence and the target, and B) affinity between the aptamer and the its complementary DNA aptamer.이 연구에서 우리는 E. coli O157에 특이적인 압타머와 상보적 DNA 압타머가 결합된 AuNP의 응집에 기초하여 E. coli O157:H7의 현장에서 신속하며 민감하게 검출할 수 있는, 압타머가 결합된 이중 기능링커 AuNP를 이용한 비색 검출법을 개발하였다. 이 시스템에서 AuNP는 이중기능링커 및 응집보조 프로브로 사용되었으며, 각 AuNP 표면은 E. coli O157:H7 대상 압타머와 상보적인 DNA 서열의 압타머로 결합되었으며, 이는 각각 이중기능링커 AuNP (BL)와 응집보조 프로브 AuNP (AA)로 명명하였다. BL은 대상균이 존재하지 않을 때에는 상보적인 서열에 의해 AA와 결합하였으며, 양적균형에 의해 특정영역(REVC, 120 – 220 μL)에서 응집 및 침전이 일어났다. 하지만, 대상균이 투입되었을 때에는 경쟁적 결합 우선순위에 의해 상보적 서열의 AA가 아니라 대상균인 E. coli O157:H7과 결합하게 된다. 이는 기존의 양적균형에서의 차이를 일으키게 되고 대조군과 유사한 응집 및 침전이 일어나기 위해서는 기존보다 더 많은 양의 BL이 반응에 참여하여야 한다. 이를 통해 대조군과 대상균 간에는 REVC 영역의 변화가 발생하게 되며, 이를 통해 대상균의 유무를 확인할 수 있었다. 검출은 가장 민감하며 구분되는 BL 영역인 100, 120, 140 μL에서 진행되었으며, 최적화된 조건에서 101 ~ 106 CFU/400 μL의 검출 범위와 57 ± 0.5 CFU/400 L의 검출 한계를 나타냈다. 또한 압타머 기반의 BL을 이용한 비색분석법은 실제 매트릭스인 코코넛워터에서도 on/off 감지 시스템으로 매우 우수하게 평가되었다. 따라서 제안된 압타머 기반의 BL을 이용한 비색분석법은 식중독을 일으키는 병원체를 현장에서 검출하는 데 적합한 방법이 될 수 있을 것으로 기대된다. 또한, 본 연구를 통해 실제 매트릭스에서 병원균 분석을 위한 앱타머 기반 비색 분석 개발에 대한 향후 연구의 기반을 제공할 수 있었다. 이 압타머 기반 비색 분석법은 A) 압타머 서열과 표적 사이의 친화성, B) 압타머와 그 상보적인 DNA 압타머 사이의 친화력이라는 두 가지 경쟁적인 상호 작용 역학에 의해 구동되었다.Ⅰ. INTRODUCTION １ Ⅱ. MATERIALS AND METHODS 5 2.1. Materials 5 2.2. Instrumentation 5 2.3. Culturing of Bacteria 6 2.4. Preparation of gold nanoparticles (AuNP) 6 2.5. Modification of AuNP with aptamer 7 2.6. Optimization of NaCl concentrations 8 2.7. Detection of E. coli O157:H7 in samples 8 2.8. Verification of the selectivity of the detection method 9 2.9. Statistical analysis 10 Ⅲ. RESULTS AND DISCUSSION 11 3.1. Overall detection procedures 11 3.2. Optimization of NaCl concentrations 17 3.3. Selectivity of the colorimetric assay 25 3.4. The range of REVC 27 3.5. Detection of E. coli O157:H7 in HEPES buffer 29 3.6. Detection of E. coli O157:H7 in coconut water 34 CONCLUSION 39 REFERENCES 41 국문초록 45석

Is the Direct Fixation of Displaced Quadrilateral Plates in Acetabular Fractures Necessary?

Quadrilateral plate fractures represent a heterogeneous group of acetabular fractures. Accurate reduction is required to prevent post-traumatic arthritis. The purpose of this study is to determine the reduction effect of the direct fixation of quadrilateral plates in acetabular fractures, and to evaluate the strength of direct fixation compared to indirect fixation. Between 2005 and 2021, 49 patients underwent surgery for open reduction and internal fixation in acetabular fractures with severely displaced quadrilateral plates. Twenty-nine patients comprised the indirect fixation group, and twenty patients comprised the direct fixation group. In a comparison of primary outcome between two groups, 10 out of 29 indirect-group patients and 1 out of 20 direct-group patients developed post-traumatic osteoarthritis, wherein the difference between the two groups is statistically significant. In the assessment of postoperative Matta\u27s radiological reduction status, 19 out of 20 patients in the direct group had achieved anatomical and congruent reduction. The treatment using a direct reduction and internal fixation improved the reduction quality of articular displacement and offered a better survivorship of the affected hip joint

Short term outcomes of topiramate monotherapy as a first-line treatment in newly diagnosed West syndrome

PurposeTo investigate the efficacy of topiramate monotherapy in West syndrome prospectively.MethodsThe study population included 28 patients (15 male and 13 female children aged 2 to 18 months) diagnosed with West syndrome. After a 2-week baseline period for documentation of the frequency of spasms, topiramate was initiated at 2 mg/kg/day. The dose was increased by 2 mg/kg every week to a maximum of 12 mg/kg/day. Clinical assessment was based on the parents' report and a neurological examination every 2 weeks for the first 2 months of treatment. The baseline electroencephalograms (EEGs) were compared with the post-treatment EEGs at 2 weeks and 1 month.ResultsWest syndrome was considered to be cryptogenic in 7 of the 28 patients and symptomatic in 21 patients. After treatment, 11 patients (39%) became spasm-free, 6 (21%) had more than 50% spasmsreduction, 3 (11%) showed less than 50% reduction, and 8 (29%) did not respond. The effective daily dose for achieving more than 50% reduction in spasm frequency, including becoming spasm-free, was found to be 5.8±1.1 mg/kg/day. Nine patients (32%) showed complete disappearance of spasms and hypsarrhythmia, and 11 (39%) showed improved EEG results. Despite adverse events (4 instances of irritability, 3 of drowsiness, and 1 of decreased feeding), no patients discontinued the medication.ConclusionTopiramate monotherapy seems to be effective and well tolerated as a first line therapy for West syndrome and is not associated with serious adverse effects

Determining the Optimal Dosage of Dexmedetomidine for Smooth Emergence in Older Patients Undergoing Spinal Surgery: A Study of 44 Cases

BACKGROUND Emergence agitation, or delirium, occurs during early recovery from general anesthesia and involves disorientation, excitation, and uncontrolled physical movements. Dexmedetomidine is an alpha agonist that has sedative, anxiolytic, analgesic, and sympatholytic activities and is used as a continuous infusion to prevent emergence agitation. This study aimed to evaluate patients aged 65 years and older undergoing general anesthesia to determine the 90% effective dose (ED90) of dexmedetomidine continuous intraoperative infusion to prevent emergence agitation. MATERIAL AND METHODS We enrolled 44 patients aged 65 years and older undergoing spinal surgery under general anesthesia. Dexmedetomidine administration commenced 30 minutes before surgery completion, with a predetermined infusion dose (μg/kg/h), without a loading dose. The initial dose was 0.2 μg/kg/h, and subsequent step size was ±0.05 μg/kg/h. We tried to find ED90 of dexmedetomidine using the biased-coin design. Vital signs, extubation quality scores, extubation-related complications, and postoperative outcomes were monitored. RESULTS Dexmedetomidine ED₉₀ for smooth emergence in older patients was 0.34 μg/kg/h. Peri-extubation vital signs remained within 20% of baseline values, without requiring pharmacological intervention. No hypoxia, hypoventilation, or post-extubation agitation occurred. In the recovery room, 1 patient briefly exhibited excitement but quickly calmed. Nine patients initially unresponsive in the recovery room fully awoke and were promptly discharged. CONCLUSIONS For older patients who are vulnerable to adverse effects of anesthetics and opioids, dexmedetomidine enables gentle awakening without adverse vital sign changes, respiratory depression, excessive sedation, or emergence agitation (ED₉₀=0.34 μg/kg/h). Further studies should involve a larger patient cohort, considering diverse medical conditions in older individuals

Association Between Obstructive Sleep Apnea and Risk of Benign Vocal Fold Lesions: A Nationwide 9-Year Follow-Up Cohort Study

Since obstructive sleep apnea (OSA) affects various parts of the body, there has been little interest about the effect of OSA on voice. The objective of this study was to evaluate the risk of benign vocal fold lesions (BVFL) in OSA patients. This study used data from the National Health Insurance Service (NHIS) database. The study group was defined as the group diagnosed with OSA between 2008 and 2011. Non-OSA groups were selected based on propensity score (PS) matching. Incidence of BVFL among participants during the follow-up was analyzed. Cox proportional hazard regression analyses were performed to evaluate the association between OSA and incident BVFL. The HR value of the OSA group calculated by considering 8 variables indicates that the risk of developing BVFL is 79% higher than that of the control group. Further, among OSA patients, patients with a history of OP had a 35% lower risk of developing BVFL. The relationships between BVFL and 7 individual variables considered were as follows: For age, HR for the 40 to 59 years group was 1.20 (95%CI, 1.09-1.32). For sex, the HR in the female group was 1.22 (95%CI, 1.10-1.35). For residential areas, the HR values for Seoul 1.39 (95%CI, 1.23-1.59). In the high economic status group, the HR was 1.10 (95%CI, 1.01-1.21). This observational study indicated that OSA is associated with an increased incidence of BVFL. The incidence of BVFL increased with older age, female sex, and high SES