恶性疟原虫质粒DNA探针在蚊媒监测中的应用研究

利用克隆的质粒型ＤＮＡ片段用光敏生物素标记后作探针．以核酸分子杂交手段．检验蚊体内的恶性疟原虫．结果表明可从多种模拟感染蚊虫标本中特异地测出恶性疟原虫感染蚊。将一组蚊研磨后集体检测时，可测出２５只蚊虫中有１只感染者或含１００个子孢子的蚊虫，这极利于大样本的快速处理．而单个蚊虫直接任于硝酸纤维素膜上检测，更适于子孢子的精确估计。既能测新鲜蚊，又能测干燥蚊标本．     

蚊体内马来丝虫幼虫的基因检测

目的：探讨蚊体内马来丝虫幼虫基因检测的新方法,并用于丝虫病监测。方法：利用基因工程技术合成马来丝虫寡核苷酸片段,经32P标记后作为探针,以斑点杂交法检测蚊体内马来丝虫幼虫。结果：该探针可从多种标本中特异地检出马来幼丝虫DNA,敏感性为2ng靶DNA量,蚊体内含1条幼虫即可出现阳性杂交。结论：该技术具有良好的实用性,可用于蚊体内马来丝虫幼虫的基因检测。     

PCR技术检测血液中和蚊体内班氏丝虫的研究

目的：寻找一种敏感、特异的班氏丝虫病的检测方法。方法：根据班氏丝虫Ssp I重复序列,合成班氏丝虫特异性引物一对;运用PCR技术检测血液中的班氏丝虫微丝蚴和蚊体内幼丝虫,并将此技术应用于原班氏丝虫流行区的疾病监测。结果：该方法可检出100μl阳性血样中的1条班氏丝虫微丝蚴;50只致倦库蚊中含有一只仅感染1条班氏幼丝虫阳性蚊亦能准确检出。马来丝虫等其它寄生虫样本的PCR结果均为阴性。用该方法检测广东省班氏丝虫监测点540份血液样本和14708只致倦库蚊样本,所有样本均为阴性。结论：该PCR反应系统灵敏度高、特异性强,可作为一种新的班氏丝虫病监测方法在现场中使用。     

PCR技术检测血液中和蚊体内班氏丝虫的研究

目的　寻找一种敏感、特异的班氏丝虫病的检测方法。方法　根据班氏丝虫SspI重复序列 ,合成班氏丝虫特异性引物一对 ;运用PCR技术检测血液中的班氏丝虫微丝蚴和蚊体内幼丝虫 ,并将此技术应用于原班氏丝虫流行区的疾病监测。结果　该方法可检出 1 0 0 μl阳性血样中的 1条班氏丝虫微丝蚴 ;5 0只致倦库蚊中含有一只仅感染 1条班氏幼丝虫阳性蚊亦能准确检出。马来丝虫等其它寄生虫样本的PCR结果均为阴性。用该方法检测广东省班氏丝虫监测点 5 4 0份血液样本和 1 4 70 8只致倦库蚊样本 ,所有样本均为阴性。结论　该PCR反应系统灵敏度高、特异性强 ,可作为一种新的班氏丝虫病监测方法在现场中使用     

班氏丝虫及帝汶丝虫对某些蚊种的实验感染

以实验室饲养的埃及伊蚊(黑眼株和雅加达株)、海滨伊蚊(台湾)、自纹伊蚊(雅加达)和野外捕集的须喙按蚊(爪哇)、常型曼蚊(雅加达)来叮吸混合感染班氏丝虫与帝汶丝虫患者进行实验。两种丝虫均能在埃及伊蚊(黑眼株)及海滨伊蚊体内发育,而帝汶丝虫     

寡核苷酸SD92探针用于蚊体内马来丝虫幼虫的检测

寡核苷酸ＳＤ９２探针用于蚊体内马来丝虫幼虫的检测山东省寄生虫病防治研究所＊（济宁２７２１３３）陈锡欣黄炳成刘克义杨宝金韩广东蚊媒体内丝虫幼虫的检测是丝虫病监测的重要内容。传统的蚊虫解剖查幼丝虫费时费力，不适应当前形势的需要。我们以合成寡核苷酸ＳＤ９２...     

淡色库蚊酯酶A2探针应用于现场抗性检测的研究

目的制备淡色库蚊酯酶A2探针,应用于现场蚊虫抗性检测。方法设计酯酶A2上下游引物,5′端地高辛标记,PCR法制备DNA探针并对扩增片段测序、分子杂交检测各现场蚊虫抗性。结果单蚊基因组DNA稀释至1/20时,抗性与敏感品系杂交点的阳性率有差别。应用酯酶A2探针检测各现场蚊虫,唐口种群的杂交阳性率为33%,微山种群的杂交阳性率为38%,滕州种群的杂交阳性率为56%。结论应用酯酶A2探针能够检测现场蚊虫抗性水平。该方法灵敏、可靠,为蚊虫有机磷杀虫剂抗性分子生物学检测提供了选择。     

上海市新江湾城淡色库蚊抗性测试结果分析

淡色库蚊是我国常见的“家蚊”,雌蚊侵入室内饱吸人血之后,多栖留在室内,完成其生殖营养周环[1]。它与人类的关系密切,不仅骚扰吸血,而且是我国虫媒病班氏丝虫的重要传播媒介,建国以来一直是除四害的主要对象。淡色库蚊是上海市的常见蚊虫,优势蚊种,也是我们日常监测的重要媒介     

温岭县基本消灭班氏丝虫病后期动态监测

本文报告1989～1992年在基本消灭班氏丝虫病的浙东南沿海地区姆坑村纵向监测结果,始监测时人群微丝蚴率为0.17%,平均微丝蚴密度为9.25条/60μl,致倦库蚊幼丝虫感染率为1.24%。1991年复查人群微丝蚴率为0.1992年蚊媒监测未发现蚊体内感染幼丝虫,结果表明监测区丝虫病的传播流行已被终止。     

乙型肝炎病毒核酸探针压电晶体传感器的初步研究

核酸探针压电晶体传感器，即由特异的单链ＤＮＡ片断与压电晶体镀膜电板交联固定制成探针，并与标本中待测的变性ＤＮＡ杂交成双链，杂交前后ＤＮＡ量的变化，可通过压电晶体振荡频率的改变判定。从含ｐＢＲ３２２－ＨＢＶ ＤＮＡ的大肠杆菌ＨＢ１０１中提取纯化重组质粒ＤＮＡ，制成乙型肝炎病毒核酸探针压电晶体传感器，检测１２例乙型肝炎感染者血清标本，结果显示５例阳性，与ＰＣＲ检测结果一致，提示其具有良好的特异性与第三     

Detection of Wuchereria bancrofti in mosquitoes by the polymerase chain reaction: a potentially useful tool for large-scale control programmes

Focally endemic bancroftian filariasis is targeted for elimination in the Nile delta of Egypt. Improved methods are needed for identifying endemic villages to be included in the control programme and for monitoring its success. We have evaluated the performance of a polymerase chain reaction (PCR) assay in estimating Wuchereria bancrofti infection in pools of Culex pipiens (1-25 females) from 2 adjacent villages with high (El Qolzom, 10.8%) and low (Kafr Shorafa, 2.1%) prevalence rates of human filariasis. This assay detects a repeated sequence in W. bancrofti deoxyribonucleic acid (DNA). Mosquitoes resting within houses were captured by aspiration and pooled by house. Houses were classified as positive or negative for human filarial infection based on night blood examinations of residents. The assay detected parasite DNA in mosquitoes from 60% of 25 infected houses and 24% of 25 uninfected houses. PCR processing of mosquitoes caught within houses of unknown filariasis infection status (44 in El Qolzom, 37 in Kafr Shorafa) identified 31.8% and 8.1% of houses, respectively, as containing infected mosquitoes. These results support the validity of the PCR assay for evaluating filarial prevalence in different villages. C. pipiens collected outdoors in dry ice-baited traps and tested by PCR (266 in Qolzom, 82 in Kafr Shorafa) did not contain parasite DNA. Pools of female mosquitoes (296 in Qolzom, 240 in Kafr Shorafa) captured in oviposition traps were also negative. We concluded that the PCR based assay is a powerful epidemiological tool that can be used for evaluating W. bancrofti infection in villages in the Nile delta and for monitoring the application of control programmes in filariasis endemic areas.     

Transmission of the nocturnal periodic strain of Wuchereria bancrofti by Culex quinquefasciatus: establishing the potential for urban filariasis in Thailand

Control programmes have reduced the prevalence of Bancroftian filariasis in Thailand to low levels. Recently, there has been an influx of more than one million Myanmar immigrants into urban centres of Thailand. The prevalence of patent Wuchereria bancrofti infection in these immigrants (2-5%) has prompted concern in the public health community that the potential now exists for a re-emergence of Bancroftian filariasis in Thailand. It is possible that an urban cycle of transmission could become established. The Myanmar immigrants are infected with the nocturnal periodic (urban) type W. bancrofti for which Culex quinquefasciatus serves as the main vector. The Thai strains of Cx. quinquefasciatus have never been reported to transmit Bancroftian filariasis. Our results of feeding experiments demonstrated that the Thai Cx. quinquefasciatus are permissive for the development of Myanmar W. bancrofti to infective third-stage larvae thus establishing the potential for establishing an urban cycle of transmission in Thailand. We also adapted the SspI repeat PCR assay for the identification of infective mosquitoes that was capable of detecting a single infective stage larvae in a pool of 100 mosquitoes.     

Surveillance and Vector Control of Lymphatic Filariasis in the Republic of Korea

ObjectivesUntil the early 2000s, lymphatic filariasis would commonly break out in the coastal areas in Korea. Through steady efforts combining investigation and treatment, filariasis was officially declared eradicated in 2008. This study surveyed the density of vector species of filariasis in past endemic areas, and inspected filariasis DNA from collected mosquitoes for protection against the reemergence of filariasis.MethodsBetween May and October 2009, mosquitoes were caught using the black night trap in past endemic coastal areas: Gyeongsangnam-do, Jeollanam-do, and Jeju-do. The collected mosquitoes were identified, and the extracted DNA from the collected vector mosquitoes was tested by polymerase chain reaction for Brugia malayi filariasis.ResultsOchletotatus togoi, Anophel es (Hyrcanus) group and Culex pipiens were most frequently caught in Jeollanam-do (Geomun Island, Bogil Island, Heuksan Island), Jeju-do (Namone-ri, Wimi-ri). and Gyeongsangnam-do (Maemul Island). DNA of B malayi was not found in Och Togoi and An (Hyrcanus) group as main vectors of filariasis.ConclusionLymphatic filariasis was not found in the vector mosquitoes collected in past endemic areas. However, considering that the proportion of vector species is quite high, there is a potential risk that filariasis could be reemerging through overseas travel or trade. Thus, there is a need to continuously monitor vector mosquitoes of lymphatic filariasis.     

Vector competence of autogenous and anautogenous Culex pipiens mosquitoes for Wuchereria bancrofti

Vector competence of autogenous and anautogenous Culex pipiens; derived from North Sinai Governorate, Egypt, for the human filaria parasite Wuchereria bancrofti was studied. After feeding on the same microfilaremic volunteers both biotypes were readily infected with the parasite (infection rates > 80%) and supported its development to the infective stage within 11 - 12 days. Infective rates of both autogenous and anautogenous mosquitoes were similar (> 95%). However, autogenous Cx. pipiens developed significantly less number of infective stage larvae (4.7 +/- 1.4 L3 / female) than did anautogenous siblings (6.7 +/- 3.6 L3 / female) (P < 0.05). Moreover, autogenous females were observed to contain twice the number of L3 larvae in the thoracic muscles and less larvae in the head region compared to autogenous counterparts. Vector competence characteristics of Cx. pipiens derived from a filariasis endemic area in Qalubiya Governorate were similar to those of anautogenous mosquitoes of North Sinai. These findings indicate that autogenous Cx. pipiens may be less efficient vector of W. bancrofti in endemic areas of Egypt.     

Some observations on filariasis in Western Samoa after mass administration of diethylcarbamazine.

An extremely efficient diethylcarbamazine administration campaign to eradicate Wuchereria bancrofti has been carried out in Western Samoa. The use of the membrane-filtration technique has shown that a large number of people exhibit extremely low microfilarial densities, often with less than 10 in 1 ml of venous blood. It was found that one of these low level microfilaria carriers readily infected the local vector Aedes polynesiensis and that development took place to the infective stage. It was estimated that 497 infective larvae of W. bancrofti will enter the human population of Western Samoa daily from these vectors. Resumption of filariasis transmission is possible and surveillance of the human and mosquito populations should be continued for a number of years and control measures taken quickly if further transmission occurs.     

Wuchereria bancrofti infection in human and mosquito populations of a Polynesian village ten years after interruption of mass chemoprophylaxis with diethylcarbamazine.

In 1991, a study on Wuchereria bancrofti microfilariae (mf) and infection rates was carried out in the human and mosquito populations of a Polynesian village where, 10 years before, the mf prevalence rate was 6.4% and twice-yearly mass treatment with 3 mg/kg of diethylcarbamazine (DEC) was interrupted. Venous blood samples were collected from 575 (97%) individuals aged 15 years or more, of whom 122 (21.4%) were mf positive. The mf carrier prevalence rate was 27.4% in males, significantly higher than that of 14% in females; it increased from 7-12% in the youngest age group (15-19 years) to 40-50% in the oldest (> or = 60 years) for both males and females. 387 mosquito collections were performed and 1748 female Aedes polynesiensis were dissected, of which 1176 were parous. Among the latter, 114 (9.7%) were infected with Wuchereria bancrofti larvae at L1, L2 or L3 stages. The mean number of larvae per mosquito was 2.46 (range 1-15). Of the 114 infected mosquitoes, 30 harboured L3 larvae, giving a 2.55% infective rate; the mean number of L3 larvae per mosquito was 1.15 (range 1-2). Such findings indicate that the interruption of systematic twice-yearly mass treatment with DEC (3 mg/kg) has resulted, after 10 years, in a substantial increase of microfilarial prevalence in humans, and in high infection rates in mosquitoes.     

Present status of bancroftian filariasis in Maceió, State of Alagoas, Brazil

Epidemiological and entomological surveys were carried out in the human and mosquito populations in Maceió, Alagoas, in order to assess the present status of bancroftian lymphatic filariasis. Examination of thick blood smears of 10,450 students from different areas of the city revealed 0.66% Wuchereria bancrofti microfilaria carriers. The distribution of filariasis is focal in the city, 80% of the individuals with patent infection living in two neighboring areas with 1.24% and 5.25% prevalence. Parallel studies performed with samples of all age groups in the human population showed similar microfilaria prevalence rates observed previously in the student survey. However, thick blood smears taken from members of families with at least one subject with patent infection gave a prevalence six times greater suggesting, increased transmission in households. The percentage of carriers was higher in the youngest age group (< 20 years). Culex quinquefasciatus mosquitos caught at the locations where the autochthonous cases were found presented natural infection rates ranging from 0.28% to 4.62%. The combination of all these findings indicates occurrence of active transmission of W. bancrofti in the urban area of Maceió, Alagoas State. Based on these data, measures for the potential control of filariasis were planned.     

Placebo-controlled community trial of four cycles of single-dose diethylcarbamazine or ivermectin against Wuchereria bancrofti infection and transmission in India

A double-blind placebo-controlled trial was carried out in 1994-98 to compare the effects of 4 cycles of single-dose diethylcarbamazine (DEC) or ivermectin on prevalence and geometric mean intensity (GMI) of microfilaraemia in the human population, infection rates in the vector population, and transmission intensity of Culex-transmitted Wuchereria bancrofti in rural areas in Tamil Nadu state, south India. Fifteen villages (population approximately 26,800) were included in the study: 5 villages each were randomly assigned to community-wide treatment with DEC or ivermectin or placebo. People over 14 kg bodyweight received DEC 6 mg/kg, ivermectin 400 micrograms/kg or a placebo, all identically packaged. After 2 cycles of treatment at a 6-month interval, the code was broken and the study continued as an open trial, with third and fourth cycles of treatment at a 12-month interval; 54-77% of eligible people (20,872) received treatment during the 4 cycles. Microfilaraemia prevalence and GMI fell by 48% and 65% with DEC and 60% and 80% with ivermectin respectively after 4 cycles of treatment. There was no change in the incidence of acute adenolymphangitis. Infection in resting mosquitoes fell significantly in all arms: 82%, 78% and 42% in the ivermectin, DEC and placebo arm, respectively. Landing mosquitoes also showed the same trend. The decline in infectivity was significant for resting (P < 0.05) and landing mosquitoes (P < 0.05) with ivermectin and DEC (P < 0.05), and for neither in the placebo group (P > 0.05). Transmission intensity was reduced by 68% with ivermectin and 63% with DEC. Transmission was apparently interrupted in 1 village with ivermectin, but infected resting mosquitoes were consistently found in this village. Single-dose community-level treatment with DEC or ivermectin is effective in reducing W. bancrofti infection in humans and mosquitoes, and may result in total interruption of transmission after several years of control. There is an immediate need to define the role of vector, parasite and community factors that influence the elimination of lymphatic filariasis, particularly the duration of treatment vis-à-vis efficacy of drugs, treatment compliance and efficiency of vectors.     

Mosquito surveillance and polymerase chain reaction detection of West Nile virus, New York State

West Nile (WN) virus was detected in the metropolitan New York City (NYC) area during the summer and fall of 1999. Sixty-two human cases, 7 fatal, were documented. The New York State Department of Health initiated a departmental effort to implement a statewide mosquito and virus surveillance system. During the 2000 arbovirus surveillance season, we collected 317,676 mosquitoes, submitted 9,952 pools for virus testing, and detected 363 WN virus-positive pools by polymerase chain reaction (PCR). Eight species of mosquitoes were found infected. Our mosquito surveillance system complemented other surveillance systems in the state to identify relative risk for human exposure to WN virus. PCR WN virus-positive mosquitoes were detected in NYC and six counties in the lower Hudson River Valley and metropolitan NYC area. Collective surveillance activities suggest that WN virus can disperse throughout the state and may impact local health jurisdictions in the state in future years.