This study was conducted to develop a real time reverse transcriptase-PCR (RT-PCR) method for the detection of viable Enterobacteriaceae in milk using primers based on the genes of ribosomal proteins S11 and S13 and to determine effects of heating and subsequent treatments on the threshold cycle (Ct) of the real time RT-PCR. Total RNA was isolated from 17 strains of bacteria including 11 strains of Enterobacteriaceae suspended in milk using a modified Tri reagent method. SYBR Green Master Mix was added to the RNA and the mixture was subjected to the real time RT-PCR. The Cts of eleven type strains of the Enterobacteriaceae in milk ($10^7$ cells) in the real time RT-PCR ranged from 21.5 to 24.6. However, the Cts of Pseudomonas fluorescens, Acinetobacter calcoaceticus, and three gram-positive bacteria were more than 40. The real time RT-PCR detected as low as $10^3$ cells in agarose gel electrophoresis. The Cts increased from 22.0 to 34.2 when milk samples contaminated with Escherichia coli ($10^7$ cells/mL) were heated at $65^{\circ}C$ for 30 min. In addition, subsequent incubation at $37^{\circ}C$ for 6 and 24 h increased the Cts further up to 36.2 and 37.2, respectively. Addition of RNase A to the bacterial suspension obtained from the heated milk and subsequent incubation at $37^{\circ}C$ for 1 h increased the Cts to more than 40. The results of this study suggests that pretreatment of bacterial cells heated in milk with RNase A before RNA extraction might enhance the ability to differentiate between viable and dead bacteria using real time RT-PCR.
Chu, Mi Ae;Jang, Yoon Young;Lee, Dong Won;Kim, Sung Hoon;Ryoo, Namhee;Park, Sunggyun;Lee, Jae Hee;Chung, Hai Lee
Clinical and Experimental Pediatrics
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v.64
no.12
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pp.652-660
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2021
Background: Viral load and shedding duration are highly associated with the transmission of severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) infection. However, limited studies have reported on viral load or shedding in children and adolescents infected with sudden acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Purpose: This study aimed to investigate the natural course of viral load in asymptomatic or mild pediatric cases. Methods: Thirty-one children (<18 years) with confirmed SARS-CoV-2 infection were hospitalized and enrolled in this study. Viral loads were evaluated in nasopharyngeal swab samples using real-time reverse transcription polymerase chain reaction (E, RdRp, N genes). cycle threshold (Ct) values were measured when patients met the clinical criteria to be released from quarantine. Results: The mean age of the patients was 9.8 years, 18 (58%) had mild disease, and 13 (42%) were asymptomatic. Most children were infected by adult family members, most commonly by their mothers. The most common symptoms were fever and sputum (26%), followed by cough and runny nose. Nine patients (29%) had a high or intermediate viral load (Ct value≤30) when they had no clinical symptoms. Viral load showed no difference between symptomatic and asymptomatic patients. Viral rebounds were found in 15 cases (48%), which contributed to prolonged viral detection. The mean duration of viral detection was 25.6 days. Viral loads were significantly lower in patients with viral rebounds than in those with no rebound (E, P=0.003; RdRp, P=0.01; N, P=0.02). Conclusion: Our study showed that many pediatric patients with coronavirus disease 2019 (COVID-19) experienced viral rebound and showed viral detection for more than 3 weeks. Further studies are needed to investigate the relationship between viral rebound and infectiousness in COVID-19.
Purpose During PET/CT examinations, the movements of internal organs caused by respiration are captured in images during multiple breathing cycles, resulting in the increases in tumor size and effects on SUV. Respiratory synchronized systems were used to evaluate tumor sizes and SUV changes. Materials and Methods Biograph mCT 64 was used for the equipment, and RPM and Anzai systems were used for the respiratory synchronized systems. We used point source and micro-phantom for an experimentation. We were performed on 12 patients who had solid tumors discovered at the base of the lung or at the top of the liver from August through September 2016. The PET images of the exhalation-to-breathing state and the CT images of the post-exhalation suspension state were gained to evaluate changes in radioactivity concentration (KBq/mL), SUVmax, cylinder diameter (mm), and tumor diameter (cm) under the conventional Static, RPM, and Anzai methods. Results The result of measuring the radioactivity concentration of the point source was RPM 94% and Anzai 91% against Static, respectively. In the two cylinders of different radioactivity in the micro-phantom, the SUVmax increased to RPM 61% and 78%, and Anzai 58% and 77% against Static, whereas the cylinder diameters decreased by RPM -26% and -28%, and Anzai -28% and -26%, each respectively. Among the patients, the SUVmax increased from a minimum of RPM 8.2% to a maximum of 94.4% against Static, and from a minimum of Anzai 7.6% to a maximum of 68.3%, respectively. As for the tumor diameters, a minimum of RPM -7.6% to a maximum of -28.9% were achieved, while the Anzai fell by a minimum of -9.6% to a maximum of -27.7%, respectively. There was no significant difference discovered in the phantom study between the RPM and Anzai, yet there was a meaningful difference in the patients' tumors (P<0.05). Conclusion The respiratory synchronized systems of RPM and Anzai yielded no significant difference in the phantom study in which the respiration was executed at regular intervals. However, it was discovered that the patients had a meaningful difference for the irregular respiratory cycle and inter-system differences. Still, the respiratory synchronized systems would be useful for the accurate diagnosis and SUV measurement as the tumor decreased in size against the existing Static and the SUV increased.
The Transactions of The Korean Institute of Electrical Engineers
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v.62
no.1
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pp.143-148
/
2013
When the fault occurs in power system, the fault-current exceeds breaking capacity of the circuit breaker. So, reliablity of the power system is decreased sharply. Superconducting fault-current limiter (SFCL) is operated without impedance in normal state. The fault-current is limited by its impedance during the fault condition. However, the SFCL has several weak points such as huge size, high-price, liquid-nitrogen operation for the real power system. In this paper, We suggested the high-speed interrupter to limit the fault-current in case of the single line-to-ground fault. In addition, we compared the high-speed interrupter with the SFCL to ensure the operation reliability. The proposed interrupter detected the fault-current through the CT, and the power was supplied by operation of the SCR control system. In this experiment, the power of high-speed interrupter was applied after the 4.8[msec] from fault instant. The on-off operation of the interrupter was started after half-cycle from the fault. The fault-current was flowed into the impedance element by the switching operation of the high-speed interrupter. So, the fault current was limited within one cycle, and then it didnt exceed the capacity of a circuit breaker. We confirmed that there was slight difference between the SFCL with high-speed interrupter in terms of limiting-time of the fault-current and switching speed of the SCR. The high-speed interrupter was considered to be more efficient than the SFCL in size, cost or reliability.
The Transactions of The Korean Institute of Electrical Engineers
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v.61
no.11
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pp.1743-1747
/
2012
The research of superconducting fault current limiter (SFCL) for reduction of the fault current is actively underway in the worldwide. In this paper, we analyzed the characteristics of a SFCL using the transformer and superconducting elements combined mutually in accordance with the fault types. The structure of this SFCL was composed of the secondary and third windings of a transformer connected to the load and the superconducting element, respectively. The provided electric power flew into the load connected to the secondary winding of the transformer in normal state. On the other hand, when the fault occurred in power system, the fault current was limited by closing the line of third winding of the transformer. At this time, the effect of the fault was minimized by opening the fault line in secondary winding of a transformer in power system. The sensing of the fault state was performed by the current transformer(CT) and then turn-on and turn-off switching behavior of the secondary line in the transformer was performed by the silicon-controlled rectifier(SCR). As a result, the proposed SFCL limited the fault current within one-cycle efficiently. Also, the degradation of the superconducting element in the normal state was avoided.
Objectives: The purpose of this study is to report the effect of combination treatment of Traditional Korean medicine (TKM) and chemotherapy on a ovarian cancer with lung metastasis patient. Methods: One ovarian cancer with lung metastasis patient was treated by TKM in conjunction with Carboplatin/paclitaxel since Feb. 2020. Repeat cycle every 3 weeks for 6 times. The patient has been treated with TKM at the same time. To evaluate the patient, symptoms were measured by Numeric Rating Scale (NRS), Eastern Cooperative Oncology Group (ECOG) and tumor size was measured by scanning with Computed Tomography (CT). Blood tests including cancer biomarker were conducted during treatment. Adverse events were evaluated by the National Cancer Institute Common Terminology Criteria for Adverse Event (NCI-CTCAE), version 5.0. Results: After treatment with Carboplatin/paclitaxel and TKM during 2 months, the size of the ovarian cancer was decreased(Partial Response, PR), size and malignant pleural effusion at right lung disappeared. And no evidence of newly developed metastatic lesions. After 2 months, the tumor response was stable disease while improving the performance and other symptoms. Conclusions: This case provides us conjunctive treatment with Conventional and Eastern medicine may have substantial benefit for patients with end-stage ovarian cancer.
Since there is no consensus about the most reliable assays to detect invasive aspergillosis from samples obtained by minimally invasive or noninvasive methods, we compared the efficacy of an enzyme-linked immunosorbent assay (ELISA) for galactomannan (GM) detection and quantitative real-time PCR assay (qRT-PCR) for the diagnosis of invasive pulmonary aspergillosis. Neutropenic, male Sprague-Dawley rats (specific pathogen free; 8 weeks old; weight, $200{\pm}20g$) were immunosuppressed with cyclophosphamide and infected with Aspergillus fumigatus intratracheally. Tissue and whole blood samples were harvested on days 1, 3, 5, and 7 post-infection and examined with GM ELISA and qRT-PCR. The A. fumigatus DNA detection sequence was detected in the following number of samples from 12 immunosuppressed, infected rats examined on the scheduled days: day 1 (0/12), day 3 (0/12), day 5 (6/12), and day 7 (8/12) post-infection. The sensitivity and specificity of the qRT-PCR assay was 29.2% and 100%, respectively. Receiver operating characteristic curve (ROC) analysis indicated a Ct (cycle threshold) cut-off value of 15.35, and the area under the curve (AUC) was 0.627. The GM assay detected antigen in sera obtained on day 1 (5/12), day 3 (9/12), day 5 (12/12), and day 7 (12/12) post-infection, and thus had a sensitivity of 79.2% and a specificity of 100%. The ROC of the GM assay indicated that the optimal Ct cut-off value was 1.40 (AUC, 0.919). The GM assay was more sensitive than the qRT-PCR assay in diagnosing invasive pulmonary aspergillosis in rats.
Kim, Jin-Young;Lee, Seung Jae;jung, Suk;Park, Min-Soo;Kang, Chun-Goo;Im, Han-Sang;Kim, Jae-Sam
The Korean Journal of Nuclear Medicine Technology
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v.19
no.2
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pp.63-67
/
2015
Purpose Among various causes that influence image quality degradation, various methods for decrease in Artifact occurred by respiration of patients are being used. Among them, this study intended to evaluate CTAC Shift correction method and additional scan compare to the Scan(Q static scan) using respiratory gated system. Materials and Methods This study was conducted on 10 patients, and used PET-CT Discovery 710 (GE Healthcare, MI, USA) and Varian's RPM system. 5.18 Mbq per kg of $^{18}F$-FDG was injected on patients, asked them to take a rest for 1 hour in the bed, and conducted test after urination. Images were visualized through Q static scan, CTAC Shift correction method, Additional scan based on the Whole body scan(WBS) with Artifact. Decrease in Artifact was compared in each image, conducted Gross Evalution, and measured changes of SUVmax. Results For image obtained through the CTAC Shift correction method through WBS with Artifact, 12~56%, Q static scan image showed 17~54% of change rate and Additional Scan showed -27~46% of change rate. In Blind Test, the CTAC Shift correction image showed the highest point with 4 points, Q static scan image showed 3.5 points, and Additional scan image showed 3.4 points. The standardized WBS scan through Oneway ANOVA and three types of Scan method showed significant difference(p<0.05), and did not show significant difference between the three Scan methods(p>0.05). However, the three Scan methods showed significant difference in Blind test. Conclusion Additional scan and Q static scan require more time than the CTAC Shift correction method, there is concern about excessive exposure to patients by CT rescan and Q static scan is difficult to apply on patients with inconsistent respiration or irregular respiration cycle due to pain. For CTAC Shift correction method, limited correction is possible and the range is limited as well. It is considered as a useful method of improving diagnostic value when hospitals use the system appropriately and develop various advantageous factors of each method.
Lee Chang Geol;Kim Joo Hang;Kim Sung Kyu;Kim Sei Kyu;Kim Gwi Eon;Suh Chang Ok
Radiation Oncology Journal
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v.20
no.2
/
pp.116-122
/
2002
Purpose : A randomized prospective study was conducted to compare the efficacy of early or late alternating schedules of radiotherapy, and carboplatin and ifosfamide chemotherapy in patients with limited-disease small cell lung cancer. Materials and Methods: From August 1993 to August 1996, a total of 44 patients with newly diagnosed, limited-disease small cell lung cancer, PS $H0\~2$, wt $loss<10\%$ were enrolled in a randomized trial which compared early alternating radiotherapy (RT)/chemotherapy (CT) and late alternating RT/CT. The CT regimen included ifosfamide $1.5\;g/m^2$ IV, d1-5 and carboplatin AUC 5/d IV, d2 peformed at 4 week intervals for a total of 6 cycles. RT (54 Gy/30 fr) was started after the first cycle of CT (early arm, N=22) or after the third cycle of CT (late arm, N=22) with a split course of treatment. Results : The pretreatment characteristics between the two arms were well balanced. The response rates in the early $(86\%)$ and late $(85\%)$ arm were similar. The median survival durations and 2-year survival rates were 15 months and $22.7\%$ in the early arm, and 17 months and $14.9\%$ in the late arm (p=0.47 by the log-rank test). The two-year progression free survival rates were $19.1\%$ in the early arm and $19.6\%$ in the late arm (p=0.52 by the log-rank test). Acute grade 3 or 4 hematologic and nonhematologic toxicities were similar between the two arms. Eighteen patients $(82\%)$ completed 6 cycles of CT in the early arm and 17 $(77\%)$ in the late arm. Four patients received less than 45 Gy of RT in the early arm and two in the late arm. There was no significant difference in the failure patterns. The local failure rate was $43\%$ in the early arm and $45\%$ in the late arm. The first site of failure was the brain in $24\%$ of the early arm patients compared to $35\%$ in the late arm (p=0.51). Conclusion : There were no statistical differences in the overall survival rate and the pattern of failure between the early and late alternating RT/CT in patients with limited-disease small cell lung cancer.
Background: Ventilator-associated pneumonia (VAP) requires prompt and appropriate treatment. Since methicillin-resistant Staphylococcus aureus (MRSA) is a frequent pathogen in VAP, rapid identification of it, is pivotal. Our aim was to evaluate the utility of quantitative polymerase chain reaction (qPCR) as a useful method for etiologic diagnoses of MRSA pneumonia. Methods: We performed qPCR for mecA, S. aureus-specific femA-SA, and S. epidermidis-specific femA-SE genes from bronchoalveolar lavage or bronchial washing samples obtained from clinically-suspected VAP. Molecular identification of MRSA was based on the presence of the mecA and femA-SA gene, with the absence of the femA-SE gene. To compensate for the experimental and clinical conditions, we spiked an internal control in the course of DNA extraction. We estimated number of colony-forming units per mL (CFU/mL) of MRSA samples through a standard curve of a serially-diluted reference MRSA strain. We compared the threshold cycle (Ct) value with the microbiologic results of MRSA. Results: We obtained the mecA gene standard curve, which showed the detection limit of the mecA gene to be 100 fg, which corresponds to a copy number of 30. We chose cut-off Ct values of 27.94 (equivalent to $1{\times}10^4$ CFU/mL) and 21.78 (equivalent to $1{\times}10^5$ CFU/mL). The sensitivity and specificity of our assay were 88.9% and 88.9% respectively, when compared with quantitative cultures. Conclusion: Our results were valuable for diagnosing and identifying pathogens involved in VAP. We believe our modified qPCR is an appropriate tool for the rapid diagnosis of clinical pathogens regarding patients in the intensive care unit.
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