Development of a loop-mediated Isothermal amplification assay for sensitive and rapid detection of Vibrio parahaemolyticus

<p>Abstract</p> <p>Background</p> <p><it>Vibrio parahaemolyticus </it>is a marine seafood-borne pathogen causing gastrointestinal disorders in humans. Thermostable direct hemolysin (TDH) and TDH-related hemolysin (TRH) are known as major virulence determinants of <it>V. parahaemolyticus</it>. Most <it>V. parahaemolyticus </it>isolates from the environment do not produce TDH or TRH. Total <it>V. parahaemolyticus </it>has been used as an indicator for control of seafood contamination toward prevention of infection. Detection of total <it>V. parahaemolyticus </it>using conventional culture- and biochemical-based assays is time-consuming and laborious, requiring more than three days. Thus, we developed a novel and highly specific loop-mediated isothermal amplification (LAMP) assay for the sensitive and rapid detection of <it>Vibrio parahaemolyticus</it>.</p> <p>Results</p> <p>The assay provided markedly more sensitive and rapid detection of <it>V. parahaemolyticus </it>strains than conventional biochemical and PCR assays. The assay correctly identified 143 <it>V. parahaemolyticus </it>strains, but did not detect 33 non-<it>parahaemolyticus Vibrio </it>and 56 non-<it>Vibrio </it>strains. Sensitivity of the LAMP assay for direct detection of <it>V. parahaemolyticus </it>in pure cultures and in spiked shrimp samples was 5.3 × 10²CFU per ml/g (2.0 CFU per reaction). The sensitivity of the LAMP assay was 10-fold more sensitive than that of the conventional PCR assay. The LAMP assay was markedly faster, requiring for amplification 13–22 min in a single colony on TCBS agar from each of 143 <it>V. parahaemolyticus </it>strains and less than 35 min in spiked shrimp samples. The LAMP assay for detection of <it>V. parahaemolyticus </it>required less than 40 min in a single colony on thiosulfate citrate bile salt sucrose (TCBS) agar and 60 min in spiked shrimp samples from the beginning of DNA extraction to final determination.</p> <p>Conclusion</p> <p>The LAMP assay is a sensitive, rapid and simple tool for the detection of <it>V. parahaemolyticus </it>and will facilitate the surveillance for control of contamination of <it>V. parahaemolyticus </it>in seafood.</p

Prediction of lowest nocturnal blood glucose level based on self-monitoring of blood glucose in Japanese patients with type 2 diabetes

Available online 13 September 2018Aims: Continuous glucose monitoring (CGM) is not available for all patients with type 2 diabetes (T2D) at risk of nocturnal hypoglycemia (NH).This study was performed to predict the lowest nocturnal blood glucose (LNBG) levels. Methods: An LNBG prediction formula was developed by multivariate analysis using the data including self-monitoring of blood glucose from a formula making (FM) group of 29 insulin-treated T2D patients with CGM. The validity of the formula was assessed by nonparametric regression analysis of actual and predicted values in a formula validation group consisting of 21 other insulin-treated patients. The clinical impact on prediction was evaluated using a Parkes error grid. Results: In the FM group with a median age of 64.0, the following formula was established: Predicted LNBG (mg/dL) = 127.4-0.836 x Age (y) + 0.119 x Self-monitored fasting blood glucose (mg/dL) + 0.717 x Basal insulin dose (U/day) (standard error of calibration 17.2 mg/dL). Based on the validation results, standard error of prediction was 31.0 mg/dL All predicted values fell within zones A (no effect on clinical action) and B (little or no effect on clinical outcome) on the grid. Conclusions: LNBG could be predicted, and may be helpful for NH prevention. (C) 2018 Elsevier Inc. All rights reserved.ArticleJOURNAL OF DIABETES AND ITS COMPLICATIONS. 32(12):1118-1123 (2018)journal articl

Sensitive and rapid detection of cholera toxin-producing Vibrio cholerae using a loop-mediated isothermal amplification

<p>Abstract</p> <p>Background</p> <p><it>Vibrio cholerae </it>is widely acknowledged as one of the most important waterborne pathogen causing gastrointestinal disorders. Cholera toxin (CT) is a major virulence determinant of <it>V. cholerae</it>. Detection of CT-producing <it>V. cholerae </it>using conventional culture-, biochemical- and immunological-based assays is time-consuming and laborious, requiring more than three days. Thus, we developed a novel and highly specific loop-mediated isothermal amplification (LAMP) assay for the sensitive and rapid detection of cholera toxin (CT)-producing <it>Vibrio cholerae</it>.</p> <p>Results</p> <p>The assay provided markedly more sensitive and rapid detection of CT-producing <it>V. cholerae </it>strains than conventional biochemical and PCR assays. The assay correctly identified 34 CT-producing <it>V. cholerae </it>strains, but did not detect 13 CT non-producing <it>V. cholerae </it>and 53 non-<it>V. cholerae </it>strains. Sensitivity of the LAMP assay for direct detection of CT-producing <it>V. cholerae </it>in spiked human feces was 7.8 × 10²CFU per g (1.4 CFU per reaction). The sensitivity of the LAMP assay was 10-fold more sensitive than that of the conventional PCR assay. The LAMP assay for detection of CT-producing <it>V. cholerae </it>required less than 35 min with a single colony on thiosulfate citrate bile salt sucrose (TCBS) agar and 70 min with human feces from the beginning of DNA extraction to final determination.</p> <p>Conclusion</p> <p>The LAMP assay is a sensitive, rapid and simple tool for the detection of CT-producing <it>V. cholerae </it>and will be useful in facilitating the early diagnosis of human <it>V. cholerae </it>infection.</p

Massive trapezial metastasis from gastric adenocarcinoma resected and reconstructed with a vascularized scapular bone graft: A case report

Rationale: Isolated metastasis to the hand bones is very rare. Only seven cases of metastasis to the trapezium have been reported. To the best of our knowledge, this is the first report of a single metastasis to the trapezium from a gastric adenocarcinoma. Patient concerns: A 62-year-old man presented with pain and massive swelling in the right carpometacarpal joint of the thumb. Diagnoses: The patient was diagnosed with trapezial metastasis of advanced gastric adenocarcinoma. Interventions: The patient underwent systemic chemotherapy with cisplatin and S-1, radiotherapy to the metastatic bone, and treatment with denosumab. One year later, the huge metastatic tumor was resected, and the hand was reconstructed using vascularized scapular bone. Outcomes: Eighteen months postoperatively, the patient was satisfied with the appearance of the reconstructed hand and was able to use his right thumb in activities of daily living. Lessons: Although rare, metastasis to the trapezium should be considered in patients with persistent and progressive thumb CMC joint pain.ArticleMedicine 96(50) : e9294(2017)journal articl

An Assessment by In Situ Hybridization Method for Pathogens of Severe Respiratory Infection

It has been shown that the in situ hybridization (ISH)method for the detection of phagocyted bacteria in polymorphonuclear neutrophils is more sensitive than conventional blood culture in patients with sepsis.The present study was designed to further evaluate the clinical utility of the ISH method, using a commercially available kit (Hybrizep(R)), for detecting pathogens in patients with severe respiratory infections. Peripheral blood was taken from patients with severe respiratory infections for both routine blood culture and the ISH method,respectively.In addition,focal samples including sputum,bronchoalveolar lavage,central and thoracic catheter, etc. were simultaneously examined for bacterial culture. A total number of 22 specimens was examined.The positive cases by ISH were 50.0% in the respiratory infections,which were significantly higher than those in blood cultures (9.1%).However, identical pathogens in the ISH method were not isolated from blood and/or other sources in respiratory sites. These findings suggest a possibility of high frequency of bacteremia or multifactorious pathogens in severe respiratory infections.The ISH method may provide additional information on serious respiratory infections for the detection of bacteremia. However, the clinical utility by the ISH method for patients with severe respiratory infection remains undetermined. Shinshu Med J 59 : 223―228, 2011Article信州医学雑誌 59(4): 223-228(2011)departmental bulletin pape

Endobronchial Argon Plasma Coagulation for the Palliation of Recurrent Tracheobronchial Adenoid Cystic Carcinoma

This is a non-final version of an article published in final form in Journal of Bronchology, 14(4):278-280, October 2007.The standard treatment of adenoid cystic carcinoma (ACC) of the airway is surgery, if possible, because survival rate is better than other treatments including radiotherapy. Although ACC shows frequent recurrence during the long-term follow-up unless there has been a complete resection (negative surgical margin), no further treatments are recommended. This report describes how argon plasma coagulation using flexible bronchoscopy has been successfully employed in the treatment of ACC after conventional therapy in 1 case of recurrence after surgery and 2 cases of inoperable patients. All of the patients are alive and healthy more than 6 years after diagnosis.ArticleJournal of Bronchology. 14(4):278-280 (2007)journal articl

Giant fetal hydrometrocolpos associated with cloacal anomaly causing postnatal respiratory distress

Persistent cloaca is a rare presentation wherein the urethra, vagina, and rectum converge into a common channel with a single perineal opening. Fetal hydrometrocolpos can result if fluid accumulates behind an obstruction of this common channel. A 29-year-old woman (G4P1021) was referred at 36 2/7 weeks of gestation for evaluation of a fetal abdominal cystic mass. Detailed ultrasonography and magnetic resonance imaging showed two symmetric cystic masses, bilateral hydronephrosis, and oligohydramnios. Elective cesarean delivery was performed at 37 0/7 weeks; the baby weighed 4043 g with Apgar scores of 5 and 6 at 1 and 5 min. Intubation was performed for respiratory distress, and the infant was noted to have an imperforate anus; persistent cloaca was diagnosed. Drainage of the hydrometrocolpos improved the infant\u27s breathing remarkably, and extubation was achieved. This child\u27s imaging findings are among the largest ever reported, and resulted in neonatal respiratory distress. © 2017 Japan Society of Obstetrics and Gynecology.Embargo Period 12 month