[目的]评价屈曲牵引下前路手术复位、减压和融合固定关节突交锁下颈椎骨折脱位的临床效果.[方法]2015年1月-2019年1月,采用屈曲牵引,前路开放复位,减压融合固定治疗关节突交锁下颈椎骨折脱位患者45例,男33例,女12例;年龄29～64岁,平均(48.6±7.2)岁.采用NDI、JOA评分、ASIA分级,以及影像检查评估临床效果.[结果]45例患者均顺利完成手术,关节突复位成功率为100.0％,手术时间平均(65.4±22.8)min,切口长度平均(6.1±0.9)cm,术中出血量平均(290.9±90.0)ml.随访时间平均(2.7±1.1)年,随时间推移,NDI和JOA评分,以及ASIA评级较均显著改善(P＜0.05),均未出现脊髓损伤加重表现.影像方面,与术前相比,术后椎间高度颈椎前凸Cobb角显著改善(P＜0.05).未发现内固定物松动、移位、断裂等现象.[结论]屈曲牵引单纯前路复位、减压融合内固定术可有效治疗合并关节突交锁的下颈椎骨折脱位.

本研究通过高通量测序技术检测了寨卡病毒(ZIKV)感染组、血餐对照组和糖水对照组埃及伊蚊中肠组织小RNA的丰度,并筛选出差异表达的6个黄病毒NIRVS和9个棒状病毒NIRVS.针对筛选出的6个NIRVS设计引物用两步法逆转录荧光定量PCR(RT-qPCR)方法验证了ZIKV感染第4、7和10天中肠组织中黄病毒NIRVS片段的表达.结果显示,黄病毒NIRVS AeFlavi53在感染第4、7天均上调,结果与小RNA测序一致,AeFlavi34B和AeFlavi4A在感染第4天上调,第10天下调,但在第7天表达无变化,与小RNA测序结果不一致,其余片段表达量在感染组与对照组间无统计学差异.RT-qPCR与高通量测序的结果一致性差,其原因可能是因为细胞中长链piRNA前体受上游转录和下游酶切多个程序的调节,针对其设计引物进行定量检测的结果不稳定.

蚊虫是疟疾、登革热、黄热病等传染病的传播媒介.蚊虫防制可以降低传染病的发生和流行.近年来研究发现,人皮肤微生物的代谢产物或产生的挥发性气味物质对蚊虫具有一定的吸引力,有望为蚊虫控制提供新方法.本文就人皮肤微生物的组成、分布及其产生的挥发性气味物质对蚊虫嗅觉行为的影响进行综述,为新型蚊虫驱避剂和引诱剂的开发及应用提供参考.

Dengue virus ( DENV ) perpetuates in an alternating cycle replication in arthropod and vertebrate hosts. It is particularly important to gain comprehensive understanding of host-virus interactions and the relationship between genetic variation and virulence. To simulate the transmit of DENN between human hosts and mosquito vectors, DENV was repeatedly passaged between human hepatocellular carcinoma cell line ( Huh-7) and mosquito cell line ( C6/36) . After 10 and 18 passages, the fitness of viral populations was measured with one-step growth curves and their virulence to mice and vector transmissibility were evaluated. Mutations in each population were determined by nucleotide sequencing. Viral fitness of DENV-2 decreased following alternating passages but increased after serial passages. Analysis of the amino acid sequences revealed that DENV that underwent continuous passage in C6/36 cells accumulated fewer mutations than virus passaged in Huh-7 cells or transferred between both cell types. Furthermore, the increased virulence that observed in serial passages might be due to the amino acid changes at positions 155 and 186 of protein E and the decreased virulence that observed in alternate passages might be due to the mutation at position 307. These results explored relationship between pathogenicity and genomic mutations of DENN and are valuable to guide the monitoring of future outbreaks.

To simulate alternative microenvirmonental habitats in vertebrates and mosquito hosts,an in vitro model was utilized to determine the gene variations'pattern of the Japanese encephalitis virus(JEV).Different JEV-SA14 mutants were obtained and compared after sequenced,and the results showed that variable loci had been found in JEV during passaging in different microenviroments.All mutations were found single-based without any insertions or deletions,which suggests that the highly conserved evolution of JEV under the natural selections during viral transmission.The fitness of JEV to different hosts evaluated were shown comparable to previous studies along with corresponding genetic mutations.The in vitro alternative passaging protocol appeared to be reliable to simulate the variant infection microenviromental habitats between vertebrated hosts and vectorial mosquitoes.

埃及伊蚊Aedes aegypti是登革病毒的主要传播媒介,埃及伊蚊感染登革Ⅱ型病毒(dengue virus typeⅡ,DENV2)后产生先天免疫反应的相关研究,有助于进一步理解埃及伊蚊感染DENV2的过程和机制.本研究通过埃及伊蚊和Aag2细胞感染DENV2进行转录组分析,与未染毒的蚊虫和细胞进行对比,筛选出12个表达变化较大的基因,之后分别构建含有这12个基因的PSL1180polyUBdsRED质粒载体,并使用细胞转染技术在Aag2细胞中建立目标基因的瞬时转染过表达模型,观察了目标基因过表达后细胞上清中DENV2的复制水平.结果显示,12个基因中羟基类固醇脱氢酶样蛋白、蜕皮激素20单加氧酶、3β-羟基类固醇脱氢酶3个基因过表达后细胞上清中病毒拷贝数出现了具有统计学意义的降低,表明这3个基因的表达可能影响DENV2在Aag2细胞中的复制.这3个基因的相关蛋白都涉及到20-羟基脱皮酮的合成,这提示20-羟基脱皮酮可能在蚊虫感染DENV2时介导了重要的免疫调节途径,为进一步研究蚊虫与虫媒病毒的互作机制提供了新的思路.

为了解海南省不同地区白纹伊蚊对常用杀虫剂的抗药性水平,为海南省城市白纹伊蚊防控提供理论依据;对海南海口、文昌和东方3个不同地区的白纹伊蚊对常用杀虫剂残杀威、毒死蜱、氯菊酯、溴氰菊酯的抗药性水平进行了测定.结果显示,白纹伊蚊对残杀威抗性倍数由1.77到3.06,对毒死蜱抗性倍数由0.81到1.42,对氯菊酯抗性倍数由2.21到3.37,对溴氰菊酯抗性倍数由3.03到5.35.结果表明,海南部分地区的白纹伊蚊对残杀威、氯菊酯、溴氰菊酯产生了一定的抗性,而对毒死蜱较敏感.

白纹伊蚊Aedes albopictus是登革病毒、寨卡病毒等多种病原体的重要传播媒介,广泛分布于我国温带和亚热带地区,以滞育卵越冬.一旦虫媒病毒扩散到我国北亚热带地区,非流行季节时期病毒的保存方式,将成为虫媒病毒研究中的重要问题.本研究开展了白纹伊蚊滞育卵的生态学研究,发现在短光照下不同地理株白纹伊蚊滞育卵的滞育率从产卵后7 d逐渐上升到21 d,至40 d趋于平缓并达到最高,滞育率在滞育前期和滞育后期统计学上差异显著.在滞育中后期,北京株白纹伊蚊滞育率相比杭州株白纹伊蚊和广州株白纹伊蚊滞育率明显较高.卵孵化后,不同地理株白纹伊蚊的卵的孵化率在滞育前期和滞育后期有显著差异.在短光照下滞育后期北京株白纹伊蚊的卵孵化率(21.45％)明显比广州株白纹伊蚊的卵孵化率(8.15％)高,提示位于温带地区的北京株白纹伊蚊较位于亚热带地区的杭州株和广州株白纹伊蚊滞育率和孵化率更高,说明位于温带地区的白纹伊蚊品系的滞育卵更能在冬季不良环境如低温等下存活,有利于增加越冬存活率.通过白纹伊蚊以滞育卵实现种群扩散、定殖的重要生物学基础,为探索新型蚊媒的控制途径提供科学依据.

埃及伊蚊和白纹伊蚊是登革热的主要传播媒介,蚊虫孳生情况的持续调查和分析,有助于对种群变化进行实时监控,从而为登革热的防治提供理论依据.2017-2020年,每年9-10月在云南省西双版纳州(勐腊县、景洪市、勐海县),临沧市(镇康县南伞镇、耿马县孟定镇),德宏州(瑞丽市)共6个市(县)调查埃及伊蚊和白纹伊蚊孳生地类型并捕捞幼虫,对4龄幼虫分类鉴定以观察两种伊蚊特征.结果 显示,此次在所调查的6个市(县)均发现两种伊蚊孳生地,共560个.孳生地类型以家用水桶类最多,占比44.82％.边境各地区居民区调查发现埃及伊蚊孳生地占比33.64％,商业区埃及伊蚊占比48.89％,两种调查地埃及伊蚊数量差异存在统计学意义,在商业区埃及伊蚊占比高于居民区.2017年西双版纳州和瑞丽市白纹伊蚊孳生地占比分别为75.51％和67.44％,2020年为36.67％和38.10％,说明两地白纹伊蚊构成比下降.两地区埃及伊蚊孳生地占比由2017年的18.37％和16.28％增长为2020年的43.33％和42.85％,说明埃及伊蚊构成比上升.结果 表明,由于埃及伊蚊比白纹伊蚊传播登革热的能力更强,2020年的西双版纳州和瑞丽市较2017年,商业区较居民区暴发登革热的潜在风险更高.

目的 研制蚊虫嗅觉风洞并通过实验验证其性能,用于驱避剂筛选和评价.方法 采用低成本、模块化的方案,设计并研制一套蚊虫嗅觉风洞,通过不同温度、湿度、风速等环境因素的调试,确定实验关键因素;通过埃及伊蚊、致倦库蚊对4种商用驱避剂涂抹人手臂和小鼠的嗅觉行为测试,验证该嗅觉风洞应用于蚊虫驱避剂筛选的可行性.结果 研制的蚊虫嗅觉风洞主要分为气路控制系统和行为筛选系统两部分;不同温度、湿度、风速等环境因素测试结果显示,温度(30±2)℃、湿度(40±3)％、风速0.2 m/s为嗅觉风洞对蚊虫嗅觉行为测试的最佳因素;未使用驱避剂时,风洞内人手臂对埃及伊蚊的引诱率为(44.1±23.1)％,小鼠对致倦库蚊的引诱率为(35.6±5.1)％,均明显高于空白对照(P＜0.05).4种商业化驱避剂对埃及伊蚊的引诱率分别为(4.4±0.4)％、(2.7±0.2)％、(3.6±1.2)％和(3.6±1.2)％,对致倦库蚊的引诱率分别为0％、0％、(3.8±3.3)％和(3.8±1.6)％,与对照组相比,使用驱避剂后人手臂和小鼠对蚊虫的引诱能力明显降低,均存在显著性差异(P＜0.05).结论 研制出一套蚊虫嗅觉风洞,通过2种蚊虫对现有商业化驱避剂的嗅觉行为测试,验证了该嗅觉风洞的性能,可用于蚊虫驱避剂的筛选和评价.

目的 通过对致倦库蚊溴氰菊酯敏感株和抗性株进行转录组分析,筛选可能与抗性相关的差异表达基因. 方法 对致倦库蚊实验室敏感株(SS株)、海南株(HN株)和溴氰菊酯药筛抗性株(RR株)3个不同抗性水平的株系进行生物测定,分别构建3个株系的cDNA文库,提取RNA,进行转录组测序.基于FPKM值进行差异性分析,对差异基因进行GO和KEGG功能注释与富集分析.筛选出致倦库蚊不同株系中表达量有显著性差异的基因,并采用实时定量PCR验证. 结果 生物测定结果显示,与致倦库蚊SS株相比,HN株和RR株的抗性倍数分别达到了4 828倍和197 241倍,为高等抗性水平.RNA-seq测序结果显示,致倦库蚊SS株、HN株和RR株转录组共注释基因转录本197 079个.GO富集分析结果显示,在结合、催化活性、细胞过程、代谢过程、单一生物过程、细胞、细胞连接、细胞器中基因富集数量较多.KEGG富集分析结果显示,在富集度最高的前20个条目中,RR株主要富集于剪接体、核糖体等通路,HN株则主要富集于剪切体、RNA转运、hedgehog信号通路、细胞周期、核苷酸切除修复、DNA复制等方向.基于FPKM值的差异性分析结果显示,致倦库蚊RR株和HN株较SS株均上调的有5 357个,均下调的有10 978个.与溴氰菊酯抗性相关基因的靶向性筛选结果显示,与SS株相比,RR株和HN株注释的细胞色素P450基因、谷胱甘肽转移酶基因、气味结合蛋白基因、气味受体基因、视觉基因、核糖体蛋白基因和碳酸酐酶基因转录本中,均上调的分别为100、17、13、4、16、191和1个,均下调的分别为253、48、43、4、15、504和9个.从上调基因中挑选的12个基因(2个细胞色素P450基因、1个谷胱甘肽转移酶基因、1个视觉基因、1个核糖体蛋白基因、1个碳酸酐酶基因、1个核酸内切酶基因、2个气味结合蛋白基因、3个气味受体基因)进行实时定量PCR验证结果显示,在RR株和HN株中,这12个基因较SS株均显著上调,其中RR株较SS株上调1.62～80.26倍,HN株较SS株上调2.01～189.48倍.相关性分析结果显示,12个目标基因的RNA-seq结果与实时定量PCR结果在HN株和RR株中均呈正相关(r=0.802、0.710).证明这12个目标基因的实时定量PCR结果与转录组测序结果趋势一致. 结论 基于高通量测序结果和定量PCR验证分析,致倦库蚊中的某些细胞色素P450基因、谷胱甘肽S转移酶基因、视觉基因、嗅觉蛋白基因可能在其对溴氰菊酯的抗性中起一定作用.

蚊虫不仅吸血骚扰,而且传播多种疾病,迄今依然是世界性的严重的公共卫生问题.近年来,新型虫媒病不断出现,传统的虫媒病死灰复燃,给蚊媒控制带来了新的挑战.随着基因编辑技术的出现与发展,特别是成簇的规律间隔短回文重复序列系统(Clustered regularly interspaced palindromic repeats-CRISPR associated sequences 9,CRISPR/Cas9)的出现,为开展蚊虫生理、生化、发育、宿主与病原体关系等诸多方面分子生物基础研究提供了靶标特异性的修饰工具,给蚊媒控制技术的发展带来了新的契机.本文主要针对CRISPR/Cas9技术在蚊媒研究领域的应用现状及进展进行综述,并探讨CRISPR/Cas9技术在蚊媒传染病防治的实际应用中所面临的问题,为蚊媒防治措施的应用及改进提供理论参考.

The phylogenetic relationships among 117 species in the tribe Aedini were analyzed and discussed based on the sequence variation of a 407 base pair (bp) fragment of the cytochrome oxidase I gene (COI gene). Phylogenetic trees were reconstructed on the basis of parsimony, maximum likelihood and Bayesian methods, and Shimodaira-Hasegawa tests were used to determine the best tree using RELL approximations. Despite differences in the taxa examined, the results were generally consistent with the recent morphological taxonomic revision on this group. The monophyly of the following genera and subgenera was corroborated: Armigeres ( subgenus Armigeres ), Bothaella, Collessius ( subgenus Alloeomyia ), Heizmannia, Jarnellius, Ochlerotatus ( subgenera Culicelsa and Woodius ), ‘Ochlerotatus’, Stegomyia ( subgenus Heteraspidion). However, the monophyly of the following genera and subgenera was not supported: Armigeres, Downsiomyia, Hulecoeteomyia, Ochlerotatus (subgenus Rusticoidus), Phagomyia, Psorophora and Stegomyia. In addition, the molecular data indicate that the genus Ochlerotatus (in part) appears to be the most primitive group and that Ochlerotatus theobaldi be a sister-species to all other taxa. Two genera, Ochlerotatus and Stegomyia, were found to be polyphyletic.

A field strain of Culex pipiens pallens collected from Beijing, China was reared and used for experimental resistance selections to pyrethroid insecticides, deltamethrin and esbiothrin. Fourth instar larvae were exposed to the concentrations of LC50 of the insecticides. After successive selections for seven generations, relative resistance increases were 12. 9-fold for deltamethrin and 4. 8-fold for esbiothrin, compared to the first population exposed to the insecticides. The deltamethrin-and esbiothrin-selected strains showed relative resistances of 217. 8 to deltamethrin and 41. 6 to esbiothrin, compared with the susceptible strains. PCR assays revealed a significant increase in knockdown resistance ( kdr) allelic frequency from F1 to F7. The kdr allelic frequencies in F7 generation of deltamethrin-and esbiothrin-selected populations were 98. 96% and 97. 87%, respectively. Strong associations were found between the frequencies of Phe1014 homozygotes, heterozygotes and the LC50 values in the bioassays. Regression analysis also revealed a significant correlation between LC50 value and the kdr allelic frequency. The study results demonstrate that high dose exposure of pyrethroid insecticides can rapidly increase the Phe1014 mutation frequency in Cx. pipiens pallens.

目的 了解淡色库蚊与骚扰库蚊触角中差异性表达的嗅觉基因,为揭示二者遗传分化特征提供参考依据. 方法 分别设置淡色库蚊吸血前、后,骚扰库蚊吸血前、后等4个实验组.每组共设4个生物学重复样品,每个样品取20只蚊虫触角.采用TRIzol法提取各组样品总RNA,采用NEBNext UltraTM RNA(R)文库构建试剂盒构建文库.对转录组基因进行测序,通过FPKM值差异性分析,筛选淡色库蚊与骚扰库蚊分别在吸血前、后表达差异显著的嗅觉基因,并进行实时荧光定量PCR (qRT-PCR)验证. 结果 淡色库蚊吸血前、后触角转录组共注释了50个气味结合蛋白(OBP)和31个气味受体(OR)基因,骚扰库蚊吸血前、后触角转录组共注释了36个OBP和12个OR.淡色库蚊吸血后较吸血前显著上调的OBP有4个,无显著下调的OBP.骚扰库蚊吸血后较吸血前显著上调和下调的OBP分别有3个.淡色库蚊吸血后较吸血前无显著上调的OR,显著下调的OR有3个.骚扰库蚊吸血后较吸血前无显著上调的OR,显著下调的OR有1个.qRT-PCR验证结果显示,淡色库蚊吸血后OBP17显著上调;骚扰库蚊吸血后OBP31、OBP44表达量显著上调,其中OBP44与测序结果一致,OBP31与测序结果相反.吸血前,骚扰库蚊OBP17显著高于淡色库蚊;吸血后,骚扰库蚊OBP31、OBP44显著高于淡色库蚊,OBP99a显著低于淡色库蚊.骚扰库蚊吸血后OR31、OR65、CPIJ001069表达量均显著上调,其中OR65和CPIJ001069表达量显著高于淡色库蚊(P＜0.05).淡色库蚊吸血后无显著变化的OR基因. 结论 淡色库蚊与骚扰库蚊的嗅觉基因OBP和OR表现出了进化分化,OBP17、OBP44可能参与调控骚扰库蚊吸血后行为.OR31、OR65、CPIJ001069可能参与调控骚扰库蚊吸血后行为.

In order to establish a rapid and sensitive detection technique for the pathogen of vector mosquito naturally infected population, this study mixed the positive samples of Culex pipiens quinquefasciatus tiretis and negative samples of uninfected encephalitis virus into mixed samples in a certain proportion, using Loop Mediated Isotherm respectively. Loop-mediated isothermal amplification (Loop Mediated Isothermal Amplification, LAMP) and real-time fluorescent quantitative PCR methods were used to detect the mixed samples of Cx pipiens quinquefasciatus infected with JEV. The results showed that when the proportion of positive samples in mixed samples was 0.01%, LAMP could still detect pathogens; The lowest detection ratio of quantitative PCR was 0.14% of the positive samples in the mixed sample. By further optimizing the concentration, reaction time and temperature of each component in the reaction system, a LAMP amplification method with high sensitivity and specificity was established.

Culex pipiens pallens is a host preference mosquito that favours birds over mammals. In present study, the odor substances of chicken Gallus gallus domesticus and pigeon Columba rupestris were collected with sorbent and feather extraction method, and the volatile components were analyzed by GC/MS method. A Y-type olfactometer was used to identify the chemicals affecting the search behavior of Cx. pipiens pallens. Sixty-nine compounds were identified from the odor substances of domestic chicken, in which fatty aldehydes, indole and p-cresol accounted for 17.4%, 10.77% and 10.67% of all the compounds. 100 compounds were identified from pigeon odor and feather extract. The main components of pigeon odor are benzoic acid (2.20%), Nonaldehyde (2.22%) and a-pinene (2.23%), and of pigeon feather extract were benzaldehyde (3.38%), cholesterol (2.38%) and myristic acid (2.18%). Two carboxylic acids (octanoic acid and myristic acid) and two aromatic compounds (indole and p-cresol) identified from chicken were found to be attractive to Cx. pipiens Pallens, while all aldehydes were found to have repellent effects. Two species of nonaldehyde and benzaldehyde identified from pigeons also have repellent effects on mosquitoes. According to the results of this study, the repellent or enticing of host odors to mosquitoes is a complex mechanism, which should be the combination of various odor substances that determines the attraction of the host to mosquitoes. In order to provide reasonable guidance for the development of attractants and repellents, some further researches are needed to confirm which odors are attractive and which have repellent effects.

1-octen-3-ol has a broad-spectrum inducing effect on the blood-sucking insects such as mosquitoes and the like. By blending the contents of 1-octen-3-ol, the field attraction effect of the attractant in different concentrations was tested in present study on the primary mosquito species in paddy field Culex tritaeniorhynchus and Anopheles sinensis, aimed to improve the attractiveness of mosquito-trapping devices to Cx. tritaeniorhynchus and An. sinensis. The results showed that 6.3%~20% of 1-octen-3-ol powder can significantly improve the capture efficiency onto the two species of mosquitoes, and 15% of 1-octen-3-ol can increase the capture effect more than 3 times, indicated that the method has good application prospect.

本文旨在建立检测登革1型和2型病毒的一步法实时荧光定量PCR方法,为登革热的诊断和预测提供技术支撑.依据登革1型和2型病毒的保守区序列设计引物和探针,建立了同时检测两个血清型登革病毒的一步法实时荧光定量PCR方法.特异性实验结果显示,该方法可检测出登1型和2型病毒,与登革3型、4型病毒、 黄热病毒、 流感H3 N2病毒均无交叉反应,具有较好的特异性.对登革1型和登革2型的灵敏度均为102 copies/μL.应用该方法,可检测出登革1型和2型病毒混合感染C6/36后不同时间点病毒复制动态.在两个血清型登革病毒混合感染C6/36细胞72～120 h登革2型病毒的拷贝数与单独感染时登革2型病毒的拷贝数相比显著下降.上述研究证实建立的一步法实时荧光定量PCR方法具有特异性好灵敏度高、 重复性好等特点,能够用于登革型和2型病毒混合感染样本的检测.

Objective To evaluate surgical treatment of chronic tibial osteomyelitis of Cierny-Mader type Ⅳ with Ilizarov technique and lesion osteotomy. Methods From January 2010 to May 2016, 39 patients with chronic tibial osteomyelitis of Cierny-Mader type Ⅳ were treated at our center. They were 33 males and 6 females, 8 to 54 years of age (average, 33.8 years). After debridement and lesion osteotomy, the tibia was fixated with Ilizarov external fixator. Bone was transported to the bone defect after corticotomy was performed on the proximal and/or distal tibial metaphyses simultaneously. Bifocal corticotomy was per-formed in 11 cases, proximal corticotomy in 21 cases, and distal corticotomy in 7 cases. The transport began 3 to 5 days after operation at a speed of 0.5 to 1.0 mm/d initially. The speed was lowered according to the bone healing and pain. Radiographic examination was done every 2 weeks to observe transporting deviation and osteogenesis in the transporting area. The transporting was adjusted whenever any abnormality was observed. The bone transporting lasted for 50 to 130 days (average, 62.4 days). Results The patients were fol-lowed up for 11 to 49 months (average, 21 months). All the soft tissue wounds healed uneventfully and there was no relapse of osteomyelitis. The bone defects in the 32 cases were reconstructed primarily. Nonunion of fracture ends happened in 5 cases and nonunion of the bone lengthening zone in 2 cases. The 7 cases of nonunion were healed after secondary bone grafting. Malalignment happened in 5 cases, 4 of which responded to timely adjustment of the external fixation and one of which had to receive secondary bone grafting after failure in adjustment of the external fixation. Ankle joint dysfunction occurred in 7 cases, 5 of which re-sponded to functional exercise and 2 of which accepted joint dysfunction because they refused surgery after unsatisfactory functional exercise. Pin tract infection of different severities occurred in 9 cases, one of which was treated by replacement of the K-wires under local anesthesia and the other 8 of which responded to rein-forced dressing change. Conclusions Chronic tibial osteomyelitis of Cierny-Mader typeⅣcan be treated by Ilizarov technique and lesion osteotomy. However, the Ilizarov technique should be improved because of the risks of multiple complications which can be reduced significantly by strengthening postoperative instruction, nursing, and regular follow-up.