疟疾四重巢式PCR检测方法的建立

目的建立一种能同时检测4种疟疾的多重巢式PCR检测方法。方法以4种疟原虫为研究对象,提取全血样本核酸,采用疟原虫18SSU r RNA作为分子标记,合成通用引物和4种疟原虫特异性引物,建立和优化四重巢式PCR反应体系。结果建立的方法在原虫之间无交叉反应,检测敏感性可达102copies/μl,经对随机选取的口岸发热样本进行检测,35份血液样本中,检出恶性疟13例、间日疟7例、卵形疟1例、以及恶性疟和间日疟的混合感染2例,阳性率为65.7%;比镜检的阳性率54.3%高,尤其是检出混合感染病例比例高,并对检测结果进行测序验证。结论建立的四重巢式PCR能同时检测4种不同疟原虫引发的疟疾,具有较高的特异性和敏感度,适合于口岸输入性疟疾的快速检测。     

一例输入性恶性疟和卵形疟混合感染病例调查

目的调查黄岩区一例输入性恶性疟和卵形疟混合感染病例,为疟疾防控提供参考。方法收集病例诊治和流行病学调查资料,采集病例抗凝血作镜检和巢式PCR检测。结果该病例在赤道几内亚务工期间有疟疾发作史和治疗史,在回国后第1天即出现寒战、发热、头痛等疟疾典型临床症状,镜检确诊为恶性疟;给予青蒿琥酯针剂和双氢青蒿素哌喹片治疗后,症状消失。此后在距上次发作的第110天又再次发病,疟疾快速诊断试纸(RDT)检测阴性,镜检误诊为间日疟;后经浙江省疾病预防控制中心巢式PCR检测,判定前后两次发病分别为恶性疟原虫和卵形疟原虫感染。结论该病例是一例输入性恶性疟和卵形疟混合感染病例,今后应加强疟疾类型的鉴别诊断能力,防止漏诊误诊。     

2015年大连市疟疾病例实验室检测结果分析

目的:分析大连市疟疾病例实验室检测结果,为疟疾诊断方法的选择提供理论依据。方法 :收集2015年大连市疟疾患者的血片和血液样本,应用镜检,RDT和巢氏PCR(Nest-PCR)分别对血片和血样进行检测。结果:17份样品镜检检出恶性疟原虫10例,卵形疟原虫1例,阴性6例;RDT检出恶性疟原虫13例,共同抗原阳性者1例,阴性3例,与镜检结果一致率为82.4%(14/17);巢式PCR检出恶性疟原虫13例,阴性4例,与镜检结果一致率为82.4%(14/17),与RDT结果的一致率为94.1%(16/17)。结论:采用镜检和RDT联合应用的方法,能够提高检测的敏感性,巢式PCR可提高虫种鉴定的准确性。     

镜检、RDT和Nest-PCR诊断疟疾的应用比较研究

目的探讨镜检、快速诊断试纸条(rapid diagnostic test,RDT)和巢式聚合酶链反应(nested polymerase chain reation,Nest-PCR)诊断疟疾应用领域及价值。方法收集2011—2015年疑似疟疾病例血样,采用镜检、RDT和NestPCR进行检测,比较分析3种方法的敏感性、特异性和综合性能。计数资料比较采用χ~2检验,P0.05有统计学意义。结果共收集疑似疟疾血样64份,镜检、RDT和Nest-PCR检测阳性率均为53.13%,3种方法恶性疟检出阳性率比较,差异无统计学意义(χ~2=0.33,P0.05)。RDT和Nest-PCR 3种诊断方法的灵敏度、特异度、阳性预测值、阴性预测值均为100.00%。结论 RDT可以应用于基层检测,但须进一步开展镜检或Nest-PCR确诊。     

1例输入性卵形疟原虫实验室检测

目的对1例输人性疑似疟疾患者的血样进行实验室确诊。方法首先制备血涂片,吉姆萨染色后镜检疟原虫;其次对血样进行RDT检测;最后利用疟原虫属特异性（通用型）和4种疟原虫种特异性的巢式PCR和多重PCR检测方法,对该血样进行分子生物学检测及分型。结合分子生物学检测结果,再对血片进行镜检复核。结果初次镜检结果为间日疟原虫,RDT结果为阴性。巢式PCR检测结果,仅扩增出预期大小约800bp的卵形疟条带;多重PCR（Pf／Pv）检测无特异性条带产生。重新对薄血膜复核镜检,改判为卵形疟原虫。结论综合巢式PCR、多重PCR、RDT和镜检等检测结果,确诊该患者为卵形疟原虫感染。     

一例输入性三日疟的实验室诊断报告

目的分析一例输入性三日疟病例的实验室检测确诊病例。方法对病例的流行病学病史作回顾分析,采集血样作血涂片镜检和疟疾快速诊断(RDT),由浙江省疾病预防控制中心通过巢式PCR检测确诊。结果该病例在本次就诊前一个月,刚从尼日利亚疟疾流行区务工返家,在国外务工期间曾有"疟疾"发病史,回国前后发病间隔2周左右。本次就诊外周血涂片镜检显示为恶性疟感染,RDT结果排除恶性疟感染,巢式PCR检测扩增出三日疟原虫条带后确诊为三日疟。结论疟疾实验室诊断需要综合判断,血涂片镜检和RDT可初步提示疟疾的类型,巢式PCR检测有助于不同疟种感染的鉴别确诊。     

1例输入性三日疟原虫病例实验室检测

目的:对1例输入性疑似疟疾患者的血样进行实验室检测。方法:首先制备血涂片,吉姆萨染色后镜检疟原虫;其次对血样进行RDT检测;最后利用疟原虫属特异性(通用型)和4种疟原虫种特异性的巢式PCR,对该血样进行分子生物学检测及分型。结果:镜检结果为三日疟原虫,RDT结果为非恶性疟感染。巢式PCR检测结果,仅扩增出预期大小约140 bp的三日疟条带。结论:综合巢式PCR、RDT和镜检等检测结果,确诊该患者为三日疟原虫感染。     

逆转录聚合酶链反应检测疟原虫混合感染的研究

〔目的〕建立在同一反应体系中能同时检测和鉴定恶性疟、间日疟的一步逆转录聚合酶链反应（RT-PCR）检测方法,并用于国境口岸归国劳务人员的疟疾检测。〔方法〕以疟原虫小亚单位核糖体核糖核酸基因为靶片段,设计恶性疟原虫和间日疟原虫的上游通用引物和各自的下游特异性引物,在同一反应体系中同时扩增恶性疟原虫和间日疟原虫的特异片段,对疟疾患者的PCR产物进行序列测定和分析。〔结果〕用建立的RT-PCR检测方法对广东口岸回国劳务人员中发现的2名疟疾患者的不同采样时间的全血进行检测,2006年10月和2007年1月采集的样本被分别扩增出预期大小为360bp和450bp特异扩增带,推测2名患者为恶性疟原虫和间日疟原虫的混合感染,其在入境时处于恶性疟的发作期和间日疟的潜伏期,均属于典型的集体输入性疟疾案例。〔结论〕RT-PCR检测疟原虫方法具有敏感性高、特异性强的特点,对国境口岸疟疾诊断、镜检质量控制和流行病学研究具有较大的实用价值。     

弥勒市1例输入性间日疟的诊断与调查分析

目的 评价综合诊断技术对非典型形态疟原虫感染病例确诊、降低输入性病例误诊或漏诊率的重要性,巩固已取得的消除疟疾成果。方法 对2023年1月16日弥勒市某医院接诊1例间日疟患者进行流行病学调查;通过镜检、抗原胶体金法(RDT)和分子生物学方法(巢式PCR)检测,确诊为1例输入性间日疟感染病例。结果 该患者血涂片镜检厚血膜查见疟原虫,薄血膜中查见配子体皱缩成团,被寄生的红细胞不胀大甚至略缩小,颜色正常与周围正常红细胞一致,与WS 259—2015《疟疾的诊断》、《疟疾防治手册》中病原学检查描述间日疟形态不同。结论 用镜检、抗原胶体金法(RDT)和巢式PCR 3种方法,对非典型形态疟原虫感染病例进行确诊至关重要。     

疟疾Dia Med Opti MAL-IT快速检测技术的临床应用

目的:探讨疟疾Dia Med Opti MAL-IT快速检测技术在临床运用的效果。方法:用疟疾Dia MedOpti MAL-IT快速检测法与疟原虫染色镜检法对144例疑似病例进行检测比较。结果:镜检法阴性92例,间日疟45例,恶性疟7例,Dia Med Opti MAL-IT快速法检测阴性94例,间日疟43例,恶性疟7例,二种检测方法差异无统计学意义(T=2,P=0.368)。结论:Opti MAL-IT快速检测法与镜检法无差别,且具有简便、快速的优点,适合对疟疾疑似病例开展临床检测。     

Evaluation of a rapid and inexpensive dipstick assay for the diagnosis of Plasmodium falciparum malaria.

Rapid, accurate and affordable methods are needed for the diagnosis of malaria. Reported here is an evaluation of a new immunochromatographic strip, the PATH Falciparum Malaria IC Strip, which is impregnated with an immobilized IgM monoclonal antibody that binds to the HRP-II antigen of Plasmodium falciparum. In contrast to other commercially available kits marketed for the rapid diagnosis of falciparum malaria, this kit should be affordable in the malaria-endemic world. Using microscopy and polymerase chain reaction (PCR)-based methods as reference standards, we compared two versions of the PATH test for the detection of P. falciparum infection in 200 febrile travellers. As determined by PCR and microscopy, 148 travellers had malaria, 50 of whom (33.8%) were infected with P. falciparum. Compared with PCR, the two versions of the PATH test had initial sensitivities of 90% and 88% and specificities of 97% and 96%, respectively, for the detection of falciparum malaria. When discrepant samples were retested blindly with a modified procedure (increased sample volume and longer washing step) the sensitivity and specificity of both kits improved to 96% and 99%, respectively. The two remaining false negatives occurred in samples with < 100 parasites per microliter of blood. The accuracy, simplicity and predicted low cost may make this test a useful diagnostic tool in malaria-endemic areas.     

套式PCR检测恶性疟原虫与间日疟原虫方法的建立及其应用研究

目的建立简便、灵敏、低成本的恶性疟原虫与间日疟原虫套式PCR检测方法,并探讨应用于间日疟原虫实验室检测的效果。方法制备抗凝静脉血的干滤纸血滴标本,采用干滤纸5%Chelex-100煮沸法提取疟原虫DNA,并进行套式PCR反应;通过比较PCR检测结果与疟原虫镜检结果,评价套式PCR检测恶性疟原虫与间日疟原虫方法的效果。结果在42份样本中,有34份血样PCR检测结果与镜检结果一致,占81%;5份镜检结果无法判断的血样,PCR检测为阳性或阴性;2份镜检结果为阴性的血样,套式PCR检测为阳性。结论套式PCR检测恶性疟原虫与间日疟原虫结果可靠,比镜检方法灵敏,有望在疟疾的实验室诊断中得到应用。     

广东省消除恶性疟后期监测和防治措施的效果评价

目的对1989年广东省消除恶性疟后期防治措施和监测效果进行评价。方法 1989～2009年广东省消除恶性疟后期所采取的防治措施和监测方法包括:疟疾病例的侦查和分类、现症病人治疗、疫点的处理、居民带虫率调查和防治措施的落实。结果广东省1989～2009年疟疾监测中,共血检12557305人,检出疟原虫阳性人数109706人,阳性率为0.87%。全省治疗有疟史病人76718人次,流行季节预防服药2381893人次,现症病人治疗96788人次,室内滞留喷洒受保护人口2400余万人次,药物浸帐保护人口934余万人次。1989年全省的疟疾发病率为0.58/万,1990～1999年平均年发病率为0.64/万,2000～2009年平均年发病率下降至仅为0.04/万,而且20年没有发现本地感染的恶性疟。结论所采取对传染源控制和传播媒介防制的方法和策略是可以阻断疟疾传播的,特别是没有发现本地感染的恶性疟病例,也没有出现输入性恶性疟的继发病例。     

2014-2018年湖北省疟疾诊断参比实验室检测结果

目的 分析2014-2018年湖北省疟疾诊断参比实验室样本检测结果,为巩固湖北省疟疾诊断水平提供科学依据.方法 收集湖北省2014-2018年疟疾网报病例诊断结果和样本,由省级疟疾诊断参比实验室通过镜检和巢式PCR对每份样本进行检测,分析不同年份和不同地区网报阳性结果和虫种符合情况,分析省级镜检和巢式PCR结果.结果 ...     

Use of rapid diagnostic tests for malaria in an emergency situation after the flood disaster in Mozambique

OBJECTIVES: To determine how diagnosis of malaria may be improved by combining the use of rapid diagnostic tests (RDTs) for Plasmodium falciparum malaria with clinical diagnosis by the presence or history of fever compared with clinical diagnosis alone in emergency situations with flood-affected displaced populations in Mozambique. STUDY DESIGN: A cross-sectional study conducted at the emergency outpatient clinic in a village in the Chòkwè district of Gaza Province, 3 weeks after Cyclone Eline hit Mozambique in February 2000. METHODS: A hundred and thirty children aged less than 15 years with clinical malaria were selected for examination by RDT and fluorescent microscopy using acridine orange as a reference test. The diagnosis of clinical malaria was made by a history of fever in the last three days or axillary temperature above 37.0 degrees C at the time of attending the emergency outpatient clinic. Two positive predictive values were calculated; RDTs combined with clinical diagnosis and clinical diagnosis alone. RESULTS: Positive predictive values of RDTs combined with clinical diagnosis by the presence of fever or history of fever were 87.6% (92/105) (95% confidence interval (CI) 80.8-92.8) compared with 74.6% (97/130) (95% CI 66.2-81.8) for clinical diagnosis alone. Five patients were diagnosed false negative. CONCLUSION: RDTs combined with clinical diagnosis has sufficient positive predictive value to be used in emergency situations, while RDTs could result in increasing failure to treat when they are used for decisions of treatment compared with clinical diagnosis alone.     

Malaria diagnosis under field conditions in the Venezuelan Amazon

To improve practical, accurate diagnosis of malaria in the Amazon rainforest of Venezuela, two rapid diagnostic tests (RDT) (OptiMAL-IT) and FalciVax) and a laboratory light microscope, used in the field with a battery-operated head lamp as an external light source, were evaluated against the standard laboratory microscope procedure for malaria detection. One hundred and thirty-six Yanomami patients were studied for the presence of malaria parasites. Thirty-three patients (24%) were positive for malaria (Plasmodium falciparum, P. vivax, P. malariae). Twenty-one (64%) of the positive patients had <100 parasites/microl. Both RDTs showed poor sensitivity (24.2% for OptiMAL-IT) and 36.4% for FalciVax) but good specificity (99% both for OptiMAL-IT) and FalciVax). Field and laboratory microscopy showed sensitivities of 94% and 91%, respectively. The kappa coefficient was 0.90, indicating a high agreement between field and laboratory microscopy. We conclude that (i) adequate slide reading cannot be substituted by either of the two RDTs in the Venezuelan Amazon and (ii) the use of a light source such as that described above makes slide reading more feasible than hitherto in remote areas without electricity.     

Comparison of Parascreen Pan/Pf, Paracheck Pf and light microscopy for detection of malaria among febrile patients, Northwest Ethiopia

Two malaria rapid diagnostic tests (RDT), Parascreen Pan/Pf^® and Paracheck Pf^®, were tested in rural health centres in Ethiopia against independent expert microscopy (the gold standard). Participants (n =1997) presented with presumptive malaria to ten health centers in Amhara Regional State during the 2007 peak malaria season (October to December). By microscopy, 475 (23.8%) suspected malaria cases were positive, of which 57.7% were P. falciparum; 24.6% P. vivax and 17.7% mixed infections. Parascreen and Paracheck were positive for 442 (22.1%) and 277 (13.9%) febrile patients, respectively. For Parascreen, P. falciparum sensitivity was 79.6%, specificity 97.4%, positive predictive value (PPV) 86.9%, and negative predictive value (NPV) 95.6%. For Parascreen, P. vivax sensitivity was 74.4%, specificity 98.6%, PPV 76.3% and NPV 98.4%. For Paracheck, P. falciparum sensitivity was 73.7%, specificity 99.2%, PPV 95.3%, NPV 94.5%. Sensitivity was significantly higher for both tests (P < 0.05) when parasite density was >100/μl of blood; in these cases Parascreen was 90.7% and 91.5% sensitive for P. falciparum and P. vivax, respectively, while Paracheck was 87.9% sensitive for P. falciparum. Parascreen thus performed adequately for both P. falciparum and P. vivax compared to expert microscopy and is more useful than Paracheck where microscopy is unavailable.     

Demonstration by the polymerase chain reaction of mixed Plasmodium falciparum and P. vivax infections undetected by conventional microscopy.

Mixed malaria infections (Plasmodium falciparum and P. vivax) are suspected to occur at a greater frequency than is detected by conventional light microscopy. To determine this frequency we carried out a prospective 'blinded' comparison of diagnosis by conventional light microscopy and enzymatic amplification of the circumsporozoite gene extracted from dried spotted blood samples. Patients were previously healthy, active duty Thai soldiers assigned to a malaria risk area presenting with malaria. Microscopy (oil immersion objective at 1000 x magnification) involved examination of Giemsa-stained thick and thin blood films by an experienced microscopist. Whole blood samples (25 microliters) dried on filter paper were used for species-specific parasite deoxyribonucleic acid (DNA) amplification by the polymerase chain reaction (PCR) and hybridization with radiolabelled P. falciparum and P. vivax probes. Of 137 consecutive cases of malaria studied, 9% (3/32) of microscopically diagnosed P. falciparum infections and 5% (5/104) of microscopically diagnosed P. vivax infections were found to be mixed by the PCR/DNA probe systems, while 1 case was diagnosed as mixed by both microscopy and PCR. The possibility that malaria patients may have undetected mixed infections should be kept in mind because of the specific therapy required both for P. falciparum and for radical cure of P. vivax.     

广东省惠州市2005—2007年疟疾流行与监测情况调查分析

目的通过对惠州市疟疾流行与监测情况调查分析，总结该市20052007年疟疾防治经验。评价该市在控制疟疾流行的防治效果。方法对20052007年疟疾疫情报告、疟疾病人个案调查、“三热”病人与流动人口的监测登记等进行流行病学分析。结果2005—2007年全市共发生疟疾42例，未发生局部暴发流行和本地恶性疟病例。结论在疟疾基本控制期间以加强基层镜检站的建设，加强流动人口疟疾管理和开展“三热”病人血检为主的监测工作，是控制疟疾传播和流行的有效措施。