发作期与间歇期间日疟原虫在不同按蚊体内发育差异的研究

目的  比较临床发作期与间歇期间日疟原虫在中华按蚊和嗜人按蚊体内发育的差异。方法在我国间日疟流行区分别采集临床发作期与间歇期间日疟病例的血样，采用体外人工膜饲感染系统在实验室同时体外人工感染中华按蚊和嗜人按蚊，在感染后7～9 d和14 d分别解剖蚊胃和唾液腺，并检测蚊体内的卵囊和子孢子数。结果临床发作期间日疟原虫感染中华按蚊的卵囊阳性率和阳性蚊比率、子孢子阳性蚊比率和感染度均低于间歇期间日疟原虫感染，临床发作期间日疟原虫感染嗜人按蚊的子孢子阳性率、阳性蚊比率和卵囊阳性蚊比率均低于间歇期间日疟原虫感染，而子孢子感染度则高于间歇期疟原虫感染。结论临床发作期与间歇期间日疟原虫体外人工感染中华按蚊、嗜人按蚊卵囊和子孢子有差异     

不同地理株嗜人按蚊对间日疟原虫的易感性

目的 比较我国不同地理株嗜人按蚊对间日疟原虫的易感性. 方法在我国间日疟流行区采集间日疟病例血样,采用体外人工膜饲感染系统在实验室同时体外人工感染广东、辽宁、江苏3个不同地理株的嗜人按蚊,在感染后的7～9 d和14 d分别解剖蚊胃和唾液腺,并检测蚊体内的卵囊和子孢子数.结果 共配对感染江苏、广东和辽宁地理株嗜人按蚊35批,感染7～9 d 3种按蚊卵囊阳性率分别为68.57%、60.00%和68.57%,感染后14 d子孢子阳性率分别为22.86%、14.29%和22.86%,3种按蚊感染卵囊和子孢子差异无统计学意义(P＞0.05).在感染后7～9 d分别解剖江苏、辽宁、广东株嗜人按蚊228、235只和228只,卵囊阳性蚊比例分别为28.07%、25.11%和26.75%.在感染后14 d分别解剖江苏、辽宁、广东株嗜人按蚊150、142只和135只,子孢子阳性蚊比例分别为10.67%、8.45%和11.85%,三者间卵囊和子孢子阳性蚊比例差异均无统计学意义(P均＞0.05).江苏、广东、辽宁株嗜人按蚊子孢子感染度差异无统计学意义(P＞0.05).结论 广东、辽宁、江苏3个不同地理株嗜人按蚊均对间日疟原虫易感.     

广东省1980—1987年疟疾流行状况

1980—1987年,疟疾年平均发病率3.65±0.40‰,较解放初期下降99.25%。年平均死亡率0.11/100万。1987年发病率≥5‰的尚有14个县(市),约450多万人口,其中发病率较高的有90余万人口。疟疾流行主要发生在海南岛少数民族地区和大陆人口流动频繁的地区。海南岛的恶性疟和混合疟病例减少,恶性疟、间日疟和三日疟的比例由抗疟前的0.623:0.365:0.012变化为0.383:0.617:0。大陆地区的恶性疟只见外地输入病例,三日疟均由输血引起,目前只见间日疟流行。1983—1987年,大陆地区共出现暴发点379个,其中336个点(88.7%)由外地传染源输入引起。在临时民工住棚解剖中华按蚊、嗜人按蚊和微小按蚊,发现子孢子阳性感染。海南岛的恶性疟原虫在体内对氯喹、喹哌、氨盼喹产生抗药性,在体外对奎宁、甲氟喹亦显示轻度抗药性。     

三峡大坝建成前影响疟疾流行的相关因素分析与监测结果

目的为了掌握疟疾疫情和按蚊密度动态以及影响疟疾流行的相关因素,为今后库区疟疾防治提供基础数据和依据。方法收集、整理三峡库区与疟疾流行的相关资料、建国后疟疾流行概况、媒介种群及分布等资料。1999年建立三峡库区疟疾监测点,监测疟疾疫情和按蚊种群、密度变化动态。结果三峡库区历史上曾有间日疟和恶性疟流行,20世纪60年代后无恶性疟发生,建国后发生过4次大流行,20世纪80年代末控制流行。传疟媒介有嗜人按蚊和中华按蚊。监测结果显示,疟疾疫情稳定,年发病控制在1/万以下;发热病人血检和儿童IFAT检测结果均为阴性;传疟媒介只有中华按蚊,5～9月叮人率在1～9.6只/人·夜之间,牛房密度在9～326只/小时人工之间。结论三峡水库蓄水前,传染源存在,媒介广泛分布,水库建成蓄水后,流速减缓、水面增宽、消落带积水均为按蚊孳生提供了有利条件,开展监测是必要的。     

广西微小按蚊和中华按蚊对海南株间日疟原虫易感性研究

广西微小按蚊中华按蚊感染海南株间日疟原虫，发现广西微小按蚊对海南株间日疟原虫易感性较高，卵囊阳性率为８５．２％，腺感染率为７０．６％，中华按蚊卵囊阳性率为１．９％，腺感染率为０；而对照组海南大劣按蚊卵囊阳性率为３８．０％，腺感染率为５６．５％。     

清流县疟疾监测结果分析

清流县位于福建省西部，辖15个乡(镇)，人口14．4万，居民大多居住在海拔200～600m之间。清流县为疟疾流行区，20世纪50、60年代间日疟、恶性疟混合流行(以间日疟为主)，主要传疟媒介为中华按蚊、嗜人按蚊和微小按蚊；70年代后仅有间日疟流行，传疟媒介为中华按蚊和嗜人按蚊。建国后曾发生3次暴发流行，经长期积级防治，于1988年达到部颁基本消灭疟疾标准。为防止疟疾发病率回升，巩固灭疟成果，从1989年起开展了疟疾监测，现将监测结果报告如下。     

嗜人按蚊、中华按蚊现场嗜血习性和传疟作用定量研究

目的观察嗜人按蚊、中华按蚊嗜血习性和传疟作用 ,为探索经济有效的疟防措施提供科学依据。方法以血检阳性病人计算发病率。人工诱饵法观察通宵叮人率。半通宵叮人率与晨间蚊帐按蚊密度之和计算睡前睡后叮人率 ,捕获新吸血按蚊 ,制成滤纸血膜送上海寄研所测定嗜血习性 ,解剖卵巢观察经产蚊比率。结果 1 .嗜人按蚊通宵叮人率平均为 3只 /人·夜 ,中华按蚊为 3 3只 /人·夜 ;睡前睡后叮人率平均分别为 2 .2 8只 /人·夜 ,4.43只 /人·夜 ,与 1 982调查比较 ,下降显著。2 .嗜人按蚊分布区 95 .0 9%的疟疾病人由嗜人按蚊传播 ,4.91 %由中华按蚊传播 ,嗜人按蚊传疟作用相当于中华按蚊的 2 0倍。 3 .嗜人按蚊媒介能量为 0 .0 75 2 ,中华按蚊为0 .0 0 60 ,基本繁殖率 <1 ;临界叮人率大于实际值。结论灭蚊、防蚊和及时治疗病人是防治疟疾的两大关键措施     

ELISA检测云南按蚊环子孢子蛋白的评价

目的　采用酶联免疫吸附测定(ELISA)技术检测疟疾媒介按蚊环子孢子蛋白(CSP),评价ELISA用于云南疟疾媒介按蚊传疟效能调查的可行性。　方法　现场捕获疟疾媒介按蚊经产蚊,每只蚊镜检3叶唾腺子孢子感染率,ELISA检测另3叶唾腺CSP;用含间日疟原虫配子体的患者血人工喂饲微小按蚊,11d后同法镜检唾腺子孢子感染率及ELISA检测唾腺CSP。在种植场捕获8种按蚊(微小按蚊、中华按蚊、多斑按蚊、迷糊按蚊、可赫按蚊、带足按蚊、菲律宾按蚊和须喙按蚊 )各龄成蚊,ELISA检测唾腺CSP。　结果　①现场捕获经产蚊1010只,镜检唾腺子孢子阳性7只,阳性率为0.69%;ELISA检测唾腺CSP阳性8只,阳性率为0.79%;其中6只蚊两项检测均为阳性,阳性率为0.59%。两法比较,差异无显著性意义(P>0.05)。②人工感染36只微小按蚊,镜检唾腺子孢子阳性27只,阳性率为75.0%;ELISA检测唾腺CSP阳性29只,阳性率为80.6%;其中有26只蚊两项检测(Pv2 10CSP)均阳性,阳性率为72.2%。两法比较,差异无显著性意义(P>0.05)。③种植场捕获8种按蚊各龄成蚊4675只,ELISA检测唾腺CSP阳性11只,阳性率为0.24%;其中Pv210阳性9只,Pf2A10阳性2只;微小按蚊、中华按蚊和多斑按蚊ELISA检测阳性     

间日疟原虫不同地理株对中华按蚊感染性的初步探讨

用上海和广西的中华按蚊感染以广西株和海南株间日疟原虫,发现两者对广西株的易感性均低,腺感染率分别为零和12.3％,而对海南株的易感性均明显较高,分别为36.4和43.6％。用2例广西间日疟患者血感染2批业已证明对当地间日疟原虫高度易感的武汉的中华按蚊,并用大劣按蚊作对照,结果2批大劣按蚊腺感染率均为100％,而2批武汉的中华按蚊腺感染率仅为4.6和30.6％。结合国内各地有关文献资料进行比较分析,认为我国间日疟原虫存在不同地理株,对中华按蚊的感染性明显不同。     

广东省当前疟疾流行特点分析

由于流动人口不断增加，传染源不断输入并在当地引起传播，广东省近年来疟疾疫情明显回升，当前疟疾流行主要有以下特点：媒介按蚊广泛存在，确认自然界有传疟作用者有微小按蚊、嗜人按蚊、中华按蚊和日月漂按蚊、前两种是主要是；广东省流行的均为间日疟，已不存在当地感染的恶性疟病人，疟疾病例主要分布在流动人口密集的珠江三角洲经济开发区的山丘地带，疟疾病人中有３／４是流动性民工。     

环介导等温扩增技术检测蚊体内间日疟原虫子孢子的研究

目的建立一种简便、快速和高敏感度的蚊体内间日疟原虫检测的环介导等温扩增方法（LAMP）。方法针对间日疟原虫环子孢子蛋白（CSP）基因种属特异性保守区域6个位点设计2对引物,以感染性按蚊、阴性按蚊、恶性疟原虫及正常人血DNA为模板评价LAMP的特异性。将间日疟原虫CSP基因质粒DNA梯度稀释,并与阴性按蚊DNA按1.0μl加1.3×10^6、1.3×10^5、1.3×10^4、1.3×10^3、1.3×10^2、1.3×10^1、1.3×10^0拷贝混合后为模板进行LAMP反应,观察其检测敏感性;将感染性按蚊与阴性按蚊DNA作1：2、1：4、1：8、1：16、1：32、1：64、1：128、1：256稀释,然后以稀释样本为模板进行LAMP反应,观察批量检测的敏感性。再用此方法与镜检解剖、巢式PCR同时检测同批次人工感染的67只按蚊,评价其应用价值。结果此法检测感染性按蚊阳性,对照组均为阴性;检测不同比例稀释的间日疟原虫CSP基因质粒DNA与按蚊DNA混合物,最低可检测1.3×10^2拷贝的间日疟原虫CSP基因质粒DNA与按蚊DNA的混合物;检测不同比例感染按蚊与阴性按蚊DNA混合样本,最低可检测出在128个按蚊中有1个感染按蚊的混合样本;用此法检测67只同批次人工感染的按蚊,检出率为47.76%,解剖镜检检出率为25.37%（χ^2=7.24,P〈0.01）,巢式PCR检出率为40.30%（χ^2=0.73,P〉0.05）。以镜检解剖作为金标准,LAMP敏感性为100%,巢式PCR敏感性为100%。结论LAMP检测蚊体内间日疟原虫简便、快速、敏感性高,具有广阔的应用前景。     

Identification of malaria species by ELISA in sporozoite and oocyst infected Anopheles from western Kenya

Enzyme-linked immunosorbent assays (ELISAs) for the circumsporozoite (CS) antigens of Plasmodium falciparum, P. malariae, and P. ovale were used to identify species of sporozoite and oocyst infections detected by dissection in Anopheles gambiae s.1. and An. funestus collected in western Kenya. ELISAs identified 92.5% of 1,113 salivary gland infections; Plasmodium species infections included 79.4% P. falciparum, 3.2% P. malariae, 1.7% P. ovale, and 2 or more Plasmodium species were detected in 15.7% of the Anopheles in which the species of parasite was identified. Identification was more likely with greater numbers of sporozoites observed in dissections, increasing from 65% ELISA positivity in mosquitoes with 1-10 sporozoites in their salivary glands to 96% in mosquitoes with over 1,000 sporozoites. ELISAs detected CS antigen in 66% of 294 Anopheles that by dissection had oocysts but uninfected salivary glands. Of 112 Anopheles with a single species of Plasmodium detected in the salivary glands, 29 (25.9%) had 1 or more additional species detected in the midgut, indicating a high potential for multiple infections. Similar proportions of Plasmodium species were found in An. gambiae s.1. and An. funestus.     

Field evaluation of an enzyme-linked immunosorbent assay (ELISA) for Plasmodium falciparum sporozoite detection in anopheline mosquitoes from Kenya

An enzyme-linked immunosorbent assay (ELISA) using a monoclonal antibody that recognizes a repetitive epitope on the circumsporozoite protein of Plasmodium falciparum was used in Kenya to assess malaria infections in Anopheles gambiae s.l. and An. funestus. The ELISA confirmed that 88% of 44 sporozoite-positive gland dissections were P. falciparum. The ELISA infection rate of 18.6% (n = 736) for individually tested mosquitoes for both species was significantly higher than the 10.4% (n = 537) salivary gland sporozoite rate determined by dissection. This difference was due to ELISA detection of medium and large sized oocysts on the midguts of infected mosquitoes which did not contain salivary gland sporozoites. From a series of 379 Anopheles that were cut at the thorax, ELISA tests on "head" and "body" portions showed that 29.5% of 95 positive mosquitoes contained circumsporozoite antigen in the body portion in the absence of salivary gland infections. This field evaluation demonstrates that the ELISA can most accurately be used to estimate sporozoite rates by cutting mosquitoes at the thorax and testing anterior portions.     

Effect of infection by Plasmodium falciparum on the melanization immune response of Anopheles gambiae

Melanization is an immune response of mosquitoes that could potentially limit Plasmodium development. That mosquitoes rarely melanize Plasmodium falciparum in natural populations might result from immuno-suppression by the parasite, as has been observed in Aedes aegypti mosquitoes infected by Plasmodium gallinaceum. We tested this possibility in Anopheles gambiae mosquitoes infected by P. falciparum by comparing the ability to melanize a Sephadex bead of infected mosquitoes, of mosquitoes that had fed on infectious blood without becoming infected, and of control mosquitoes fed on uninfected blood. Rather than being immuno-suppressed, infected mosquitoes tended to have a stronger melanization response than mosquitoes in which the infection failed and than control mosquitoes, possibly because of immune activation after previous exposure to invading parasites. This finding suggests that P. falciparum relies on immune evasion rather than immuno-suppression to avoid being melanized and confirms that natural malaria transmission systems differ from laboratory models of mosquito-Plasmodium interactions.     

Estimate of Plasmodium falciparum sporozoite content of Anopheles stephensi used to challenge human volunteers

Plasmodium falciparum infected Anopheles stephensi, taken from a group of mosquitoes which had been used to challenge recipients of (NANP)3-TT vaccine, were tested for P. falciparum sporozoite content by an immunoradiometric assay. Seventy-six percent were infected with mean and median sporozoite equivalents per mosquito of 220,994 and 217,398, respectively (SD = 54,911). This sporozoite density is greater than that usually found in the field. These data suggest that this challenge for evaluating P. falciparum sporozoite vaccines is a demanding test of immunity.     

A retrospective examination of mosquito infection on humans infected with Plasmodium falciparum

A retrospective examination was made of archival data collected between 1940 and 1963 on the infection of mosquitoes with Plasmodium falciparum. Patients were undergoing malariatherapy for the treatment of neurosyphilis. A total of 913 lots of Anopheles quadrimaculatus and An. albimanus were fed on 173 patients. Mosquito infection continued to occur in a few patients beyond 200 days of patent parasitemia. The primary period of mosquito infection occurred during the first 20 days of gametocytemia. Of the 311 lots of mosquitoes fed during this period, 209 (67.20%) were infected, and of these, 163 had greater than 50% of the mosquitoes in the lots infected with at least one oocyst. During secondary periods of gametocytemia, 293 (78.76%) of 372 lots of mosquitoes were infected. The highest percentages of mosquitoes were infected from four days before to four days following peak gametocyte density. Mosquito infection rates were similar to those seen in studies with splenectomized Aotus monkeys experimentally infected with P. falciparum.     

Plasmodium coatneyi: observations on periodicity, mosquito infection, and transmission to Macaca mulatta monkeys.

Plasmodium coatneyi has adapted well to experimental studies with Macaca mulatta monkeys and Anopheles dirus mosquitoes. Studies were made to determine 1) the course of asexual parasitemia, 2) periods when infective gametocytes were produced, 3) the laboratory-reared mosquitoes susceptible to infection, 4) the mosquito most capable of transmitting the infection to monkeys via bite, 5) the pattern of recrudescence, and 6) the prepatent periods following the bites of infected An. dirus mosquitoes. The period when infective gametocytes are produced is concentrated primarily in the first week when parasitemia exceeds 1,000/microl. Mosquitoes were more heavily infected on days when the asexual parasite counts were highest. Gametocyte counts were generally low. Mature forms of the parasite markedly sequestered giving a pattern of high-low periodicity. Anopheles dirus and An. freeborni mosquitoes were nearly equal in terms of their ability to support oocyst development. Other species (An. stephensi, An. maculatus, and An. gambiae.) were less supportive. High sporozoite densities in the salivary glands were frequently produced in An. dirus and sporozoite transmission was obtained via the bites of these mosquitoes after 12-18 days of extrinsic incubation. Prepatent periods ranged from 10 to 15 days. The presence of frequent parasitic recrudescences suggests mechanisms similar to that seen in human infections with P. falciparum. It is proposed that P. coatneyi in M. mulatta monkeys can be a suitable model for studies on cerebral pathology, vaccine efficacy, and the testing of antimalarial drugs.     

荧光定量PCR用于按蚊体内疟原虫子孢子检测的研究

目的 建立一种用于按蚊体内疟原虫子孢子定量检测和虫种鉴别的荧光定量PCR方法。方法 采用针对4种人体疟原虫18S rRNA基因属特异性保守区的1对引物, 以疟原虫18S rRNA基因重组质粒与按蚊DNA的混合物为模板, 进行反应体系和反应条件优化, 验证方法的特异性, 并通过熔解曲线分析进行虫种鉴别。应用阴性按蚊DNA稀释的间日疟原虫18S rRNA基因重组质粒为模板制作标准曲线, 并分别以质粒DNA和实验室子孢子感染阳性的按蚊DNA为模板分析建立的荧光定量PCR方法的敏感性。在反应体系中加入不同部位、 不同用量按蚊DNA, 以探讨按蚊DNA对检测效果的影响。结果 该方法对按蚊、 人血DNA均无扩增, 对4种疟原虫DNA均有特异性扩增且扩增产物的熔解温度 （Tm） 易于区分, 三日疟原虫、 恶性疟原虫、 卵形疟原虫和间日疟原虫的Tm值分别为71.0、 72.7、 73.9 ℃和75.9 ℃。标准曲线中循环阈值（Ct值） 与模板浓度具有良好的相关性 （相关系数r = -0.99）。最低可以检出含50拷贝的质粒DNA或32倍稀释的子孢子阳性按蚊DNA样本。按蚊DNA对荧光定量PCR反应具有抑制作用。荧光定量PCR的Ct值在实验内和实验间均具有良好的重现性。结论 新建立的SYBR Green I染料荧光定量PCR方法具有较高的敏感性和特异性, 可用于按蚊体内疟原虫子孢子的检测, 并可同时对4种人体疟原虫进行鉴别。     

Role of the prevalent Anopheles species in the transmission of Plasmodium falciparum and P. vivax in Assam state, north-eastern India

In north-eastern India, Anopheles minimus, An. dirus and An. fluviatilis are considered the three major vectors of the parasites causing human malaria. The role in transmission of the other Anopheles species present in this region is not, however, very clear. To examine the vectorial role of the more common anopheline mosquitoes, the heads and thoraces of 4126 female Anopheles belonging to 16 species (collected using miniature light traps set in human dwellings in a foothill village in the Jorhat district of Assam state) were tested, in ELISA, for the circumsporozoite proteins (CSP) of Plasmodium falciparum or the VK-210 and VK-247 polymorphs of P. vivax. Sixty-five pools of head-thorax homogenates, representing 10 different species of Anopheles, were found reactive, giving an overall minimum prevalence of infection (MPI) of 1.58%, with a 95% confidence interval (CI) of 1.21%-2.0%. Among the CSP-reactive pools of mosquitoes, 31% were positive only for P. falciparum, 45% only for P. vivax VK 247, 6% only for P. vivax VK 210, and 18% for both P. falciparum and P. vivax VK 247. The results indicate that not only the proven vector, An. minimus s.l. (MPI = 0.71%), but also several species of Anopheles previously considered unimportant in the epidemiology of malaria, especially An. aconitus (MPI = 3.95%), An. annularis (MPI = 5.8%), the An. hyrcanus group (MPI = 0.48%), An. kochi (MPI = 1.28%), the An. philippinensis-nivipes complex (MPI = 0.94%), and An. vagus (MPI = 3.87%), are important vectors in the foothills of Assam.