云南省4种伊蚊的乙型脑炎病毒分离物的研究

在云南省西南边境9县市捕获伊蚊属雌性成蚊16种19367只,用细胞法和乳鼠法分离病毒.从185批6491只白纹伊蚊中分离到病毒2株,从50批1605只剌扰伊蚊中分离到病毒2株,从23批772只窄翅伊蚊中分离到病毒2株,从4批103只阿萨姆伊蚊中分离到病毒1株.其它12种共10396只伊蚊的病毒分离物为阴性.分离到的7株病毒经免疫荧光、酶免疫、血凝抑制和中和试验鉴定,均为乙型脑炎病毒(JE virus).白纹伊蚊是野外竹林的优势蚊种.分析认为白纹伊蚊在当地乙型脑炎病毒保存和传播中起重要作用,刺扰伊蚊、窄翅伊蚊和阿萨姆伊蚊亦可参与该病毒的传播.     

白纹伊蚊垂直传播乙型脑炎病毒的研究

用５株乙型脑炎病毒（ＪＥｖｉｒｕｓ）经口感染白纹伊蚊,证明该蚊能感染和传播乙型脑炎病毒,对感染雌蚊的１和子２代卵３４４３粒,幼虫７０２１只和成虫４８５３只,分１７１批检查乙型脑炎病毒,子１代的批阳性率,卵为４６．６７％（１４／３０）,幼虫为２１．２１％（７／３３）。雌性成虫为２３．７８％（９／３８）,雄性成虫为１５．００％（３／２０）;子２代幼虫,雌性和雄性成虫和批阳性率依次为２３．５３％（４／１     

登革热的传播模式、蚊媒及防控

在自然界存在两种登革热传播模式:人-伊蚊-人循环,蚊媒是埃及伊蚊与白纹伊蚊。猴-伊蚊-猴循环,蚊媒是白纹伊蚊与白雪伊蚊群。我国学者首先于1975年从无输入性病例的我国西南边疆山林地区的白纹伊蚊体内分离到登革热病毒4型,白纹伊蚊承担两种传播模式的中介。本研究介绍了埃及伊蚊与白纹伊蚊的生态习性与全球及在中国的分布。认为在我国厦门地区迄今为止还未曾发现过埃及伊蚊的存在,也简介了沃尔巴克体新技术防控蚊媒研究的进展。     

白纹伊蚊和埃及伊蚊对基孔肯雅病毒的易感性和传播性的研究

用４株基孔肯雅病毒经口感染白纹伊蚊和埃及伊蚊,进行了易感性和传播性的研究。结果表明,这两种蚊虫对基孔肯雅病毒易感。无论白纹伊蚊或埃及伊蚊,感染后第５－６天即可通过吸血将病毒传播给乳鼠,至第８－１３天,传播率可高达５５．５５％－１００％。感染蚊亦可经叮咬将病毒传播给小鸡。埃及伊蚊的易感性和传播率高于白纹伊蚊。实验还发现,不同来源毒株之间存在一定差异,如分离自云南白纹伊蚊的Ｍ８１株的感染率和传播率均高于其它毒株。这些结果表明,白纹伊蚊和埃及伊蚊在基孔肯雅病毒的保存和传播中起重要作用。     

福建省白纹伊蚊中登革病毒的分离与鉴定

近年来福建省登革热(Dengue fever,DF)输入性病例持续存在,且登革热的主要传播媒介白纹伊蚊在全省内广泛分布,为了解福建省福州市登革热的媒介白纹伊蚊携带登革病毒(Dengue virus,DENV)状况,2017年10月7日在福州市台江区元一花园小区内开展伊蚊监测,采用双层叠帐法捕获255只白纹伊蚊蚊体研磨液上清提取核酸后用实时荧光RT-PCR法检测DENV特异性核酸,将检测阳性的蚊体研磨液上清接种C6/36细胞进行病毒分离,成功分离到1株DENV病毒株mosquito13/Fujian/2017;经实时荧光RT-PCR法鉴定所分离病毒株的血清型为I型;利用型特异性引物通过RT-PCR扩增病毒E基因并测序进行分子遗传特性分析;E基因核苷酸和氨基酸同源性分析显示,该毒株与2017年10月17日同小区本地登革热病例血清中分离得到的登革毒株E基因序列完全一致,与越南2014年分离株KT825033/Vietnam/2014核苷酸(99.7%)和氨基酸(99.8%)同源性最高;系统进化树分析表明所分离登革病毒毒株的基因型为I型,与东南亚地区的越南,泰国,柬埔寨等国家进化关系相近,可能输入来源于东南亚国家。本研究证实了登革热外潜伏期的存在以及白纹伊蚊在登革热疫情传播过程中的媒介作用,提示在登革热的防控工作中媒介登革病毒监测、检测的重要性,也提示福建省需要加强输入来源监测,特别是东南亚入境人员的监测。     

豫南罗索线虫防制轮胎积水内白纹伊蚊的试验

白纹伊蚊Aedes albopictus是我国和东南亚地区最常见的蚊种之一,分布面广,与人关系密切,为乙型脑炎和登革热的传播媒介。防制白纹伊蚊对消除此两种病的流行和传播具有重要意义。汽车的废旧轮胎多堆放于露天场所,雨后积水成为白纹伊蚊适宜的孳生地,附近居民深受其害,为此选择了某厂区轮胎积水环境投放豫南罗索线虫(Rommanomermis yunanensis)作     

用免疫荧光法检测登革病毒抗原

建立了为检测登革病毒抗原的直接免疫荧光法。将标记荧光素的抗IV型登革病毒的免疫腹水IgG抗体,滴加到感染了该病毒原型株的新生小白鼠脑印片和白纹伊蚊细胞系盖玻片培养上进行结合,都观察到特异的荧光。而未经感染或感染了其它型别登革病毒或日本乙型脑炎病毒的新生小自鼠脑印片上,均未观察到特异的荧光。这种特异的荧光能被未标记的特异性抗体阻断或抑制。在吸过感染了该病毒的小白鼠血的饲养的部份雌性白纹伊蚊头压片上,观察到特异的荧光,而在未吸过感染血的雌性白纹伊蚊头压片上,都未观察到特异的荧光。在流行区捕得的21只雌性白纹伊蚊中8只的头压片和82只中5只的涎腺、胃和卵巢压片中的细胞浆内观察到IV型登革病毒抗原的特异性荧光。而用此法对非流行区饲养的雌性白纹伊蚊的各种压片中都未观察到特异荧光。     

中国白纹伊蚊亚组伊蚊的研究——Ⅰ.成蚊

白纹伊蚊亚组(Aedes albopictus subgroup)是复蚊亚属(Stegomyia)伊蚊的重要类群之一,例如其中的白纹伊蚊是我国登革热的媒介之一,而且在福建、四川等地,它被认为与乙型脑炎的传播有关。本亚组所属种类,尤其是它们的幼虫和蛹,都很近似,因而过去有些种类不易区分。最近黄耀民(Huang,1972a;1979)对东南亚的本亚组蚊种作了比较详细的分类研究,才澄清了一些问题,提高了这方面的水平。     

云南白纹伊蚊细胞株对登革热、基孔肯亚病毒敏感性及增殖规律的观察

自Singh建立蚊细胞以来,为虫媒病毒的研究提供了新方法。singh又进行了白纹伊蚊和埃及伊蚊细胞株的比较,认为白纹伊蚊细胞更敏感,应用于虫媒病毒的分离得到满意的结果。我所1978年建立了一株白纹伊蚊细胞株,对登革病毒增殖较好。1979年我们从当地扑获蚊虫,建立了一株云南白纹伊蚊细胞株(ACY7),对登革、基孔肯亚病毒进行敏感性及增殖规律的观察,现将结果报告如下:     

白纹伊蚊抗溴氰菊酯品系的筛选及其对登革病毒的易感性

【目的】建立白纹伊蚊Aedes albopictus抗溴氰菊酯的蚊虫品系,对比白纹伊蚊敏感株和抗性株对登革病毒的易感性差异。【方法】采用浸渍法测定溴氰菊酯对白纹伊蚊幼虫的半数致死浓度(LC_(50));再以LC_(50)水平的溴氰菊酯对白纹伊蚊幼虫进行群体筛选至第11代,并通过接触筒法检测各代成蚊抗性。以白纹伊蚊敏感株和获得的抗性株(第9代)雌蚊吸食含2型登革病毒(DENV-2)的血餐,于感染后0,4,7和10 d解剖蚊虫,收集中肠、卵巢和唾液腺,通过RT-PCR和实时荧光定量PCR分别检测各组织的DENV-2病毒感染率和感染量。【结果】白纹伊蚊经溴氰菊酯筛选至第9代后抗性趋于稳定。第9代抗性株幼虫的LC_(50)为0.053 mg/L,抗性倍数为10.58,成蚊生测的蚊虫死亡率为80%,已达到中度抗性。感染后0 d,所有蚊虫的中肠均可测得DENV-2,而且抗性株的平均病毒感染量高于敏感株;后续时间点敏感株与抗性株蚊虫中肠均保持92.75%~97.18%的病毒感染率,且二株之间无显著性差异(P0.05)。感染后4 d,两株蚊虫的卵巢中均可检测到DENV-2,感染后7 d和10 d的卵巢病毒感染率均显著高于4 d时(P0.05),但在7 d和10 d两个时间点,敏感株和抗性株之间均无统计学差异(P0.05),然而抗性株平均病毒感染量在每一个时间点都显著高于敏感株(P0.05)。蚊虫唾液腺于感染后7 d检测到DENV-2,10 d时唾液腺的病毒感染率无明显升高且两株蚊虫之间病毒感染率和感染量均无统计学差异(P0.05)。【结论】经溴氰菊酯的筛选使白纹伊蚊幼虫及成蚊抗性水平逐渐升高,建立了白纹伊蚊抗溴氰菊酯的实验室品系。DENV-2对白纹伊蚊抗性株和敏感株各个组织的感染率相近,但感染量有所不同,说明对溴氰菊酯有中度抗性的白蚊伊蚊对登革病毒的易感性在一定程度上发生了改变。     

Mosquito vectors of the 1998-1999 outbreak of Rift Valley Fever and other arboviruses (Bagaza, Sanar, Wesselsbron and West Nile) in Mauritania and Senegal

Following an outbreak of Rift Valley fever (RVF) in south-eastern Mauritania during 1998, entomological investigations were conducted for 2 years in the affected parts of Senegal and Mauritania, spanning the Sénégal River basin. A total of 92 787 mosquitoes (Diptera: Culicidae), belonging to 10 genera and 41 species, were captured in light traps. In Senegal, Culex poicilipes (41%) and Mansonia uniformis (39%) were the most abundant species caught, whereas Aedes vexans (77%) and Cx. poicilipes (15%) predominated in Mauritania. RVF virus was isolated from 63 pools of Cx. poicilipes: 36 from Senegal in 1998 and 27 from Mauritania in 1999. These results are the first field evidence of Cx. poicilipes naturally infected with RVFV, and the first isolations of this virus from mosquitoes in Mauritania - the main West African epidemic and epizootic area. Additional arbovirus isolates comprised 25 strains of Bagaza (BAG) from Aedes fowleri, Culex neavei and Cx. poicilipes; 67 Sanar (ArD 66707) from Cx. poicilipes; 51 Wesselsbron (WSL) from Ae. vexans and 30 strains of West Nile (WN) from Ma. uniformis, showing differential specific virus-vector associations in the circulation activity of these five arboviruses.     

First record in America of Aedes albopictus naturally infected with dengue virus during the 1995 outbreak at Reynosa, Mexico

Abstract Mosquito collections were conducted during a dengue outbreak in Reynosa, Tamaulipas, Mexico, July-December 1995. A total of 6694 adult mosquitoes (four genera and nine species) were captured, of which 2986 (78.3% females and 21.7% males) were Aedes albopictus and 2339 (39.7% females and 60.3% males) were Ae.aegypti. These two species comprised 84.2% of the total collection. Specimens were grouped into pools, nearly 50% of them processed for detection of virus by cythopathic effect in C6-36 and VERO cell cultures and by haemagglutination test. Five pools gave positive haemagglutin-ation reactions and were examined by immunofluorescence using monoclonal antibodies to flavivirus and to dengue virus. One pool of ten Ae.albopictus males was positive for dengue virus: serotypes 2 and 3 were identified by serotype-specific monoclonal antibodies arid confirmed by RT-PCR. This is the first report of Ae.albopictus naturally infected with dengue virus in America. Also, it is the very first time Ae.albopictus males have been found infected with dengue virus in the wild.     

Growth characteristics of the veterinary vaccine candidate ChimeriVax-West Nile (WN) virus in Aedes and Culex mosquitoes

In 1999 West Nile (WN) virus was introduced to North America where this flavivirus has spread rapidly among wildlife (especially birds) transmitted by various species of mosquitoes (Diptera: Culicidae). Increasing numbers of cases and deaths among humans, horses and other domestic animals require development of effective vaccines. 'ChimeriVax-West Nile(vet)' is being developed for use as a veterinary vaccine to protect against WN infection. This chimeric virus contains the pre-membrane (prM) and envelope (E) genes from the wild-type WN NY99 virus (isolated from a flamingo in New York zoo during the 1999 WN epidemic) in the backbone of yellow fever (YF) 17D vaccine virus. Replication kinetics of ChimeriVax-WN(vet) virus were evaluated in mosquito cell culture (Aedes albopictus C6/36), in WN vector mosquitoes [Culex tritaeniorhynchus Giles, Cx. nigripalpus Theobald and Cx. quinquefasciatus Say (Diptera: Culicidae)] and in YF vectors [Aedes aegypti (L) and Ae. albopictus (Skuse)], to determine whether these mosquitoes become infected through feeding on a viraemic vaccine, and their potential infectivity to transmit the virus. Growth of ChimeriVax-WN(vet) virus was found to be restricted in mosquitoes, compared to WN virus in Ae. albopictus C6/36 cells. When inoculated intrathoracically, ChimeriVax-WN(vet) and YF 17D viruses did not replicate in Cx. tritaeniorhynchus or Cx. nigripalpus; replication was very restricted compared to the wild-type WN virus in Cx. quinquefasciatus, Ae. aegypti and Ae. albopictus. When fed on hanging drops with ChimeriVax-WN(vet) virus (7.7 log10 PFU/mL), none of the Culex mosquitoes became infected; one Ae. albopictus and 10% of the Ae. aegypti became infected, but the titre was very low and virus did not disseminate to head tissue. ChimeriVax-WN(vet) virus had a replication profile similar to that of the attenuated vaccine virus YF 17D, which is not transmitted by mosquitoes. These results suggest that the natural mosquito vectors of WN and YF viruses, which may incidentally take a bloodmeal from a vaccinated host, will not become infected with ChimeriVax-WN(vet) virus.     

Are Aedes albopictus or other mosquito species from northern Italy competent to sustain new arboviral outbreaks?

The Asian tiger mosquito Aedes albopictus (Skuse) (Diptera: Culicidae), native to Southeast Asia, has extended its geographical distribution to invade new temperate and tropical regions. This species was introduced in 1990 to Italy and has since become the main pest in urban settings. It was incriminated as a principal vector in the first European outbreak of chikungunya virus (CHIKV) in the province of Ravenna (Italy) in 2007. This outbreak was associated with CHIKV E1-226V, efficiently transmitted by Ae. albopictus . The occurrence of this outbreak in a temperate country led us to estimate the potential of Ae. albopictus to transmit CHIKV and dengue virus (DENV), and to determine the susceptibility to CHIKV of other mosquito species collected in northern Italy. Experimental infections showed that Ae. albopictus exhibited high disseminated infection rates for CHIKV (75.0% in Alessandria; 90.3% in San Lazzaro) and low disseminated infection rates for DENV-2 (14.3% in San Lazzaro; 38.5% in Alessandria). Moreover, Ae. albopictus was able to attain a high level of viral replication, with CHIKV detectable in the salivary glands at day 2 after infection. In addition, the other three mosquito species, Anopheles maculipennis Meigen, Aedes vexans vexans (Meigen) and Culex pipiens L., showed variable susceptibilities to infection with CHIKV, of 0%, 7.7% and 0–33%, respectively. This information on vector competence is crucial in assessing the risk for an outbreak of CHIKV or DENV in Italy.     

Isolation of the spirochaete Borrelia afzelii from the mosquito Aedes vexans in the Czech Republic

During the years 1993–1995, a total of 3580 culicine mosquitoes of six species were collected in South Moravia, Czech Republic, and examined by dark-field microscopy for the presence of borreliae. Females of Aedes cantans , Ae. sticticus , Ae. vexans , Culex pipiens and Cx pipiens biotype molestus (but not Ae. geniculatus or Culiseta annulata ) harboured spirochaetes, the frequencies ranging from 0.7% to 7.8%. One isolate (BR-53) from Ae. vexans was identified as Borrelia afzelii genospecies. The potential role of mosquitoes in the epidemiology of Lyme borreliosis should be investigated.     

Flaviviral disease mosquito vector surveillance in Incheon Metropolitan City and the Hwaseong area,Gyeonggi‐Do,Republic of Korea,in 2015

Owing to global climate change, the global resurgence of vector‐borne infectious diseases and their potential to inflict widespread casualties among human populations has emerged as a pivotal burden on public health systems. In this study, the prevalence of flaviviral diseases transmitted by mosquitoes, and their target vector diversity, abundance, and distribution was investigated to enable the mapping of hotspots for these diseases. For the surveillance of the vector mosquitoes carrying flaviviruses during April to November 2015, female mosquitoes were collected to study whether they carried pathogens from abroad at seven locations in Incheon Metropolitan City (Incheon) as a typical urban area and Hwaseong‐si (= city, Hwaseong) of Gyeonggi‐do (= province) as a rural area. A total of 15 species belonging to seven genera (29,102 female mosquitoes) were collected with black‐light and BG‐Sentinel? traps at a collection rate of 260 per trap/night from whole collection locations. The most collected mosquito species in Incheon were Aedes vexans nipponii (species ratio (SR), 29.9%) and the Culex pipiens complex (SR, 28.8%), followed by Anopheles sinensis s.l. (SR, 27.9%) and Ochlerotatus koreicus (SR, 7.1%). From the results of viral RNA detection, five flaviviruses were found in 20,981 individuals (excluding An. sinensis; 696 pools) in the Cx. pipiens complex and Ae. vexans nipponii. Three Japanese encephalitis virus (JEV)‐positive pools were from the Cx. pipiens complex, a Chaoyang virus pool was found from Ae. vexans nipponii, and the remaining unidentified flavivirus pool was from Cx. pipiens. The three JEV‐positive pools were phylogenetically grouped as genotype V. The results of our study demonstrate that enhanced monitoring and long‐term surveillance of these vector viruses are of great public health importance.