Determining the Efficacy of CRISPR-Cas9 Mechanisms on the uppP Gene of Klebsiella pneumonia using Lysozyme Assay

@#Introduction: Multidrug resistance bacteria is alarming worldwide. A lot of research were done and are ongoing to search for the best, convenient and economically affordable ways to fight them. With the latest genome editing tool; Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) technology, this research was performed to develop a novel strategy to genetically modify the genome and inhibit the growth of Klebsiella pneumoniae (UPM ESBLKP1), an Extended Spectrum Beta Lactamases (ESBL) organism. Methods: A CRISPR-Cas9 vector was constructed together with guide RNAs designed specifically for the targeted uppP gene, a gene responsible for bacterial cell growth and protection. Results: The growth and cell wall integrity of the modified Klebsiella pneumoniae (ΔUPM ESBLKP1) were significantly inhibited and reduced, respectively. Interestingly, wild type Klebsiella pneumoniae showed a normal growth curve while modified strains showed a faster doubling rate when supplemented with Luria-Bertani media. In contrast, slower growth rate of modified strain was observed in the M9 minimal media. This explained the higher doubling rate of mutants on nutrient rich medium earlier is being related to gene recovery. They grew slowly in the minimal media as they were adapting to a new environment while recovering the uppP gene and surviving, proving the success of its gene modification. Conclusion: The developed CRISPR-gRNA system was able to modify the targeted Klebsiella pneumoniae gene hence providing an opportunity to develop a new drug for Klebsiella pneumoniae infection as an alternative to antibiotics.

Experiencia en pacientes operados de roncopatía y apnea obstructiva del sueño / Clinical experience in patients with surgery of rhonchopathy and obstructive sleep apnea syndrome

RESUMEN Introducción: La uvulopalatofaringoplastía (UPPP) es un procedimiento ampliamente difundido en el tratamiento del ronquido y el síndrome de apnea e hipopnea obstructiva del sueño (SAHOS), ya que permite aumentar el área de sección transversal de la vía aérea superior y eliminar tejidos obstructivos. Conocer el grado de satisfacción de los pacientes con la cirugía es importante para nuestro desempeño. Objetivo: Evaluar la satisfacción de los pacientes con el procedimiento UPPP con cirugía nasal con datos subjetivos mediante la aplicación de una encuesta para ser respondida de forma anónima. Material y método: Estudio observacional, descriptivo. Se envió vía email una encuesta de 4 preguntas cerradas creada en la plataforma online MonkeySurvey a los pacientes que fueron sometidos a UPPP (faringoplastía de relocalización) con cirugía nasal entre 2015 y 2016. Resultados: 27 pacientes respondieron la encuesta. Sesenta y seis coma seis por ciento tenían ronquido primario y/o SAHOS leve, 33,3% SAHOS severo. La edad media al momento de la cirugía fue 41 años. El tiempo de seguimiento medio fue 10 meses (324 meses). Noventa y dos coma cinco por ciento de los pacientes están satisfechos con la cirugía UPPP; 81,4% refieren que recomendarían la cirugía a otra persona. Noventa y dos coma cinco por ciento refieren mejoría en los ronquidos. Noventa y cinco coma seis por ciento de los pacientes con somnolencia diurna refiere mejoría. Conclusión: La cirugía UPPP con técnica faringoplastía de relocalización combinada con cirugía nasal en pacientes con ronquido primario y SAHOS ha demostrado una alta tasa de satisfacción según la percepción de los pacientes en el seguimiento desde los 3 meses hasta los 2 años posoperatorios.

ABSTRACT Introduction: Uvulopalatopharyngoplasty (UPPP) is a widely used surgical procedure for snoring and obstructive sleep apnea syndrome (OSAS), since it allows to increase of the cross-sectional area of the upper airway and elimination of obstructive tissues. Knowing the degree of satisfaction of patients with surgery is important for our performance. Aim: To evaluate patient satisfaction with UPPP procedure (relocation pharyngoplasty) with nasal surgery with subjective data through the application of a survey to be answered anonymously. Material and method: Observational, descriptive study. A survey of 4 closed questions created in the MonkeySurvey online platform was sent to patients who underwent UPPP with nasal surgery between 2015 and 2016 via email. Results: 27 patients answered the survey. 66.6% had primary snoring and/or mild OSAS, 33.3% severe OSAS. The average age at the time of surgery was 41 years. The mean follow-up time was 10 months (3-24 months). 92.5% of patients are satisfied with UPPP surgery; 81.4% reported that they would recommend surgery to another person. 92.5% reported improvement in snoring. 95.6% of patients with daytime somnolence reported improvement. Conclusion: UPPP surgery with relocation pharyngoplasty technique combined with nasal surgery in patients with primary snoring and OSAS has shown a high satisfaction rate according to the perception of patients at follow-up from 3 months to 2 years postoperative.

The Modified Uvulopalatopharyngoplasty(UPPP)to Prevent the Complications and Destroy of UPPP / 医学研究杂志

Objective To improre typieal proeedure of UPPP,increase clinical effects and deerease complioations caused by the operation.Methods Twenty eight patients with obstructive sleep apnea syndrome who had been confirmed by PSG in our hospital from 2003 to 2005 were involved in this study.1)improving stripping method:the fat and tonsil were removed by electric knife.2)improving molding method:the muscle of uvula was maitained,but the fat was removed;mucous and the tissue under it were double-layer sutured.3) the data before and 6 months after UPPP were analyzed by questionnaire and polysomnography(PSG)detection.Result No fissuration occurred.The distance between soft palate and posterior wall was 3?0.6 before operation and it was 6?0.8 6 months after operation.The effect is 100% based on the questionnaire investigation and PSG detection.Conclusions 1)Hemostasis of electric knife was reliable;2)Double-layer suture of mucous and the tissue under it can avoid hemorrhage and insure molding of pharyngeal cavity;3)The results of the uvula preservation can improve the effect of the UPPP by increasing the nasopharyngeal and oropharyngeal cavity,and avoiding velopalatal insufficiency.

Laser-assisted uvulopalatoplasty and laser uvulopalatopharyngoplasty for snoring and obsrtuctive sleep apnea: Long-term postoperative patient's satisfaction / 대한이비인후과학회지

BACKGROUND AND OBJECTIVES: Laser-assisted uvulopalatoplasty (LAUP) and laser uvulopalatopharyngoplasty (Laser-UPPP) are well established and highly successful operation in the treatment of snoring and obstructive sleep apnea (OSA). But, most reports are based on short-term follow-up results. This study was performed to evaluate the patient's satisfaction rate after long-term postoperative follow-up, and to investigate prognostic factors of good surgical outcome, time of recurrence, and sequelae. MATERIALS AND METHOD: 102 patients were operated by LAUP (n=77) and Laser-UPPP (n=25) between Jul,1993 and Dec,1996. These patients were followed up more than 2 years (mean 3.1 years). The degree of snoring and apnea related symptoms were evaluated using questionnaires preoperatively and postoperatively. RESULTS: In 95 (93%) of 102 patients, symptoms were satisfationally reduced in 6 months of surgery. After 2 years the success rate dropped to 68% (70 of 102 patients. In the dissatisfied group 32 patients, most failures occurred between 6 and 12 months after surgery. Prognostic factors of good surgical outcomes were lower preoperative body mass index value, absence of postoperative weight gain, and combined nasal surgery. There was no major postoperative complication, but temporary sequalae ocurred as nasal regurgitation (8.8%), hypernasality (5.9%), pharyngeal foreign body sensation (13.7%), postoperative bleeding (6.9%), and long-lasting pain over 3 weeks (12.7%). CONCLUSION: Long-term follow-up more than 18 months is mandatory to evaluate postoperative results of LAUP and Laser-UPPP. To achieve good postoperative outcomes in the treatment of snoring or OSA, perioperative weight reduction should be recommended, and nasal surgery should be considered in patients with nasal obstruction.

Analysis on Outcomes and Predict of Laser Uvulopalatopharyngoplasty / 대한이비인후과학회지

BACKGROUND: Laser uvulopalatopharyngoplasty(Laser UPPP) is a modified operative procedure of UPPP using laser for the patient of obstructive sleep apnea syndrome. It is based on the progressive enlargement of the airspace in the oropharynx, to eliminate or reduce obstruction that occurs during sleep, by successive vaporization of the soft palate, wide posterior tonsil pillars and redundant posterior mucosa and resection of hypertrophied palatal tonsils. OBJECTIVES: The objectives of this study were to evaluate the success rate of the operation and to access the value of predictors of outcome in patients with obstructive sleep apnea syndrome. MATERIALS AND METHODS: Laser UPPP was performed in 23 patients with obstructive sleep apnea syndrome based on preoperative polysomnography and preoperative and postoperative polysomnography were compared to evaluate the success rate of operation. Each patient underwent preoperative fiberoptic nasopharyngoscopy with Mueller maneuver. RESULTS: 73.9% of patients had greater than 50% reduction in apnea index and the improvement of snoring was obtained in 69.5% of patients. No polysomnography parameter could accurately predict the changes in sleep after laser UPPP. There was no significant difference between responder and nonresponder group concerning the body mass index(BMI). But there was a significant difference between two groups in severity of obstruction during Mueller maneuver. CONCLUSION: The results of this study suggest that laser UPPP may be effective treatment for the obstructive sleep apnea syndrome with a high success rate and Mueller maneuver would be beneficial in selecting the candidates for laser UPPP by its high predictive efficacy.

Clinical Efficacy of Uvulopalatopharyngoplasty in the Treatment of Obstructive Sleep Apnea Syndrome / 결핵

BACKGROUND: Uvulopalatopharyngoplasty(UPPP) has become the most common surgical treatment for obstructive sleep apnea syndrome(OSAS). However, the results of this therapeutic modality have been quite variable with successful results by several authors and poor results by others. Until recently, in Korea, there is only a few reports about the clinical efficacy of UPPP. A prospective study was undertaken to evaluate the effectiveness and complications of UPPP. METHODS: Twenty-six OSAS patients who had undergone UPPP with preoperative and postoperative polysomnographic studies were included in this study. Two definitions of surgical success were used. The responder was defined, using a conventional criteria, as a 50% or more reduction in apnea index(AI) or apnea-hypopnea index(AHI) after UPPP, or a postoperative Al of <10 or AHI of <20. The initial cure was defined, using our own criteria, as a postoperative Al of <5 or AHI of <10. Complications were categorized in two groups early(disorders during the first 10 postoperative days) and late. RESULTS: Eighteen patients(69.2%) were responders, and ten patients(38.5%) were considered as initial cure. On the other hand, in five patients(19.2%), postoperative polysomnographic data demonstrated deterioration compared with preoperative data. Reduction rate of Al or AHI following UPPP was not significantly related to the preoperative body mass index, Al or AHI. There was no significant change of sleep architecture before and after UPPP in responder and initial cure groups. Early complications such as pain, dyspnea, bleeding, nasal reflux, dysphagia or wound disruption were observed in all patients. Late complications such as nasal reflux, voice change, dysphagia, loss of taste, pharyngeal dryness or foreign body sensation were discovered in 22 patients(84.6%). However, all early and late complications were of minor importance. CONCLUISON: The response to UPPP was favorable in approximately 70% of OSAS patients. However, the initial cure rate of UPPP was relatively low. We suggest that selection of more appropriate surgical candidates and adequate surgical protocol is necessary to obtain a more successful result with UPPP.

Risk of anesthesia operation for obstructive sleep apnea syndrome / 重庆医科大学学报

Objective:To explore the way of diminishing perioperative mortality of patient with obstructive sleep apnea syndrome(OSAS).Methods:The death reason of a patient with OSAS was analyzed and relevant articles were reviewed.Results:Asphyxia death mostly took place during anesthesia induction and convalescence as patients experienced general anesthesia for descending uranostaphyloplasty(uppp).Conclusion:The risk of anesthesia for the patient with OSAS must be fully recognized.Sedation amnesia and slow induction in pernasal intubation was an effective way for the operation.