Fate of a bacteriophage in Aedes aegypti, Anopheles quadrimaculatus (Diptera: Culicidae), and Periplaneta americana (Orthoptera: Blattidae)

The viability of a bacteriophage of Escherichia coli was unaffected by injection into the hemocoel of the mosquitoes, Aedes aegypti and Anopheles quadrimaculatus, but was reduced by injection into the cockroach, Periplaneta americana. Treatment of the cockroach with India ink, known to be phagocytized in the cockroach hemocoel, did not block the reduction of phage viability. Phage viability was unaffected by incubation with gut homogenates of A. aegypti but was possibly affected by homogenates of P. americana.     

Isolation and characterization of polymorphic microsatellite markers from Asian malaria mosquito Anopheles sinensis (Diptera: Culicidae)

Microsatellite-containing region were isolated and characterized in Anopheles sinensis, a primary vector of malaria parasites in Asia. An enrichment protocol yielded 252 microsatellite sequences. We designed primers to amplify 20 unique microsatellites, 14 of which amplify cleanly and were polymorphic. A survey of 24 individuals showed that 12 loci are highly variable with the number of alleles ranging from two to 11, and expected heterozygosity ranging from 0.116 to 0.903. These markers will be useful for population genetic studies and genome mapping in A. sinensis.     

Behavioural and electrophysiological responses of the malaria mosquito Anopheles gambiae Giles sensu stricto (Diptera: Culicidae) to human skin emanations

Behavioural and electrophysiological responses of Anopheles gambiae Giles sensu stricto (Diptera: Culicidae) to human skin emanations collected on glass beads were studied using a dual-port olfactometer and electroantannography. Glass beads to which skin emanations from human hands had been transferred elicited a level of attraction similar to a human hand. The attractiveness of these handled glass beads faded away 4 h after transfer onto the beads. Storage at -20 degrees C for up to 8 weeks showed a decreased but still attractive effect of the beads. In a choice test between one individual and four others, the emanations from the reference individual were significantly more attractive in three out of four cases. The headspace of handled glass beads elicited a dose-dependent EAG response. The substances causing EAG activity could be removed partially by dichloromethane, ethanol and pentane-ether. Glass beads provide a suitable neutral substrate for the transfer of human odour to enable chemical analysis of the human skin emanations for identification of kairomones of anthropophilic mosquitoes.     

尼日利亚南部阿贝奥库塔地区冈比亚按蚊复合体的形态特征研究（双翅目：蚊科）

对采自尼日利亚南部城市阿贝奥库塔冈比亚按蚊复合体Anopheles gambiae complex的形态特征进行了研究。依据2005年8月至2006年7月灯诱捕获的364个冈比亚按蚊复合体标本,分别对它们的触角、翅、喙、前足、中足和后足6个部位的长度进行了测量,对月平均值进行回归分析,同时利用差异系数（co-efficient of difference,CD）进行近缘分析。分析显示,各特征的长度平均值雨季大于旱季,但是回归分析表明长度变化与季节不显著相关（P>0.05）。差异系数分析结果表明,仅触角长度和翅长显示此复合体为两个不同的种群(CD>1.28),而其他特征值表明为同一种群。因此,该研究结果提示触角长度及翅长对冈比亚按蚊复合体近缘种的区分有重要参考价值。     

斯氏按蚊肽聚糖识别蛋白基因PGRP-LC1的克隆及功能分析

天生免疫系统是昆虫抵御外界病原入侵的主要方式。目前研究发现, Imd信号通路与按蚊感染柏氏疟原虫Plasmodium berghei的强度密切相关, 而PGRP-LC1是Imd信号通路最上游的受体之一。为了研究斯氏按蚊Anopheles stephensi肽聚糖识别蛋白PGRP-LC1, 采用RT-PCR并结合RACE技术克隆斯氏按蚊PGRP-LC1基因, 通过序列比较分析, 得到两条cDNA序列, 其开放阅读框分别为1 365 bp和1 290 bp, 3′非编码区为320 bp, 5′非编码区为240 bp。将两条cDNA分别命名为AsPGRP-LC1a（GenBank注册号 GU214232）和AsPGRP-LC1b（GenBank注册号GU214233）。AsPGRP-LC1a编码454个氨基酸, 分子量约为49.07 kDa;AsPGRP-LC1b编码429个氨基酸, 分子量约为46.3 kDa。AsPGRP-LC1b比AsPGRP-LC1a少一个长度为75 bp的外显子, 该外显子在冈比亚按蚊Anopheles gambiae PGRP-LC1基因的某些可变剪切形式中也有发现。分别将两个斯氏按蚊PGRP-LC1基因在冈比亚按蚊细胞系L3-5和斯氏按蚊细胞系MSQ43中过量表达, 通过双荧光素酶检测系统检测抗菌肽的表达情况, 结果显示克隆得到的PGRP-LC1基因在两种细胞系中均能够启动Imd信号通路, 为进一步研究斯氏按蚊的Imd信号通路提供了依据。     

Impact of permethrin-treated bednets on malaria transmission by the Anopheles gambiae complex in The Gambia

Malaria vector mosquitoes belonging to the Anopheles gambiae complex were studied in four hamlets in The Gambia. All inhabitants were given bednets treated either with a placebo (milk) in two hamlets or with the pyrethroid insecticide permethrin (500 mg/m2) in two other hamlets. Malaria transmission occurred mainly during a few weeks of the rainy season, in September and October 1987. The indoor resting densities of mosquitoes in permethrin-treated hamlets were reduced, and we estimated over 90% reduction in biting on man by An. gambiae Giles sensu stricto in these hamlets. No mosquitoes were found under permethrin-treated bednets compared with eighty-one recovered from placebo-treated bednets. Mosquitoes exited more readily from rooms where permethrin-treated bednets were used than from rooms with placebo-treated nets. The annual mean probability that a child would receive an infective bite was estimated to be 0.09 in hamlets with insecticide-treated bednets, compared with 1.9 where placebo-treated bednets were used. Permethrin-treated bednets are therefore recommended as a means of effectively reducing the risk of exposure to malaria transmission, particularly in areas of low seasonal transmission.     

林氏按蚊线粒体全基因组序列的测定及基于线粒体基因组的按蚊属系统发育分析(英文)

【目的】对林氏按蚊Anopheles lindesayi完整的线粒体基因组进行测序及分析,依据已知的线粒体基因组构建并讨论按蚊属蚊虫的分子系统发育关系。【方法】对林氏按蚊线粒体基因组进行测序、注释,并对其基本特征和基本组成进行分析。基于串联的13个蛋白质编码基因的核苷酸序列和氨基酸序列,用ML法和贝叶斯法构建林氏按蚊和按蚊属其他32种蚊虫的系统发育树,据此探讨按蚊属蚊虫的系统发育关系和系统分类。【结果】林氏按蚊线粒体基因组全长为15 366 bp,包含13个蛋白质编码基因,22个tRNA基因,2个rRNA基因和一段控制区。林氏按蚊线粒体基因组呈现明显的AT偏斜和GC偏斜,AT偏斜为正,GC偏斜为负。除了COX1使用TCG和ND5使用GTG作为起始密码子以外,其他蛋白质编码基因的起始密码子均遵循ATN原则;终止密码子为TAA或者T。除了tRNA^Ser(AGN)以外,其他的tRNA基因均呈现典型的三叶草二级结构。控制区AT含量最高,为94.54%。滑窗分析显示蛋白质编码基因是用于构建亚属或属水平系统发育关系的最佳分子标记。系统发育树强烈支持塞蚊亚属Cellia、按蚊亚属Anopheles、徕蚊亚属Nyssorhynchus和柯特蚊亚属Kerteszia均为单系群。小五斑按蚊An. atroparvus和四斑按蚊An. quadrimaculatus A这两个种聚到一起,从传统的形态分类上讲,它们和林氏按蚊均属于按蚊亚属按蚊系蚊虫。但本研究构建的4个系统发育树均显示,(小五斑按蚊An. atroparvus+四斑按蚊An. quadrimaculatus A)和林氏按蚊被属于迈蚊系的中华按蚊分开,这为两个系的分类提供了新的论点。【结论】本研究获得了林氏按蚊的完整的线粒体基因组,探析了按蚊属的线粒体基因组特征和系统发育关系,为进一步研究蚊科线粒体基因组和系统发育关系提供了依据。     

Modification of matrix polysaccharides during avocado (Persea americana) fruit ripening: an assessment of the role of Cx-cellulase

The role of C_x-cellulase (EC 3.2.1.4) in fruit ripening and softening is unknown. In the present study, avocado ( Persea americana ) fruit, a rich source of C_x-cellulase, were examined to determine if the enzyme plays a role in ripening-related hemicellulose metabolism. Hemicelluloses (4 M alkali-soluble) from avocado fruit exhibited a very broad distribution of polymer sizes and an overall decrease in M_r during ripening. Polymers affected were primarily those of large M_r (relative molecular mass). The characteristic total hemicellulose M_r distribution and changes with ripening were also evident for xyloglucan (XG), a putative substrate for avocado C_x-cellulase. Hydrolytic activity toward hemicelluloses from preripe fruit was detected in crude buffer-soluble protein extracts derived from ripe avocado mesocarp tissue. XG was also degraded, and in a pattern similar to that observed during ripening. Purified C_x-cellulase also exhibited activity against specific components of isolated hemicelluloses; however, in contrast to the crude protein. C_x-cellulase alone was without influence on the M_r distribution of avocado XG. Protein depleted of C_x-cellulase was capable of moderate XG depolymerization. We conclude from the present studies that the enzyme C_x-cellulase is not involved in the ripening-related depolymerization of XG in avocado fruit.     

The importance of avocado (Persea americana Mill.) fruits anthracnose and factors influencing the disease in Mana district,south-western Ethiopia

Anthracnose disease surveys were conducted in 25 farmers’ orchards, wholesaler and retailer shops in south-west Ethiopia. In addition, harvesting and postharvest practices, and storage conditions influencing disease development were studied with observation and questionnaire. The assessment results indicated significant variation among farmers’ orchards with the highest incidence (84.0 ± 16.7%) and severity index (26.0 ± 5.4%). Anthracnose damage of fruit was higher at retailers (76.7 ± 20.8%) than in the wholesalers shop (56.7 ± 32.5%). The total number of isolates identified was 249 and Colletotrichum gloeosporioides was the predominant pathogen proved by pathogenecity test. Among the major factors, harvesting avocado fruits with children (88%) and climbing on the tree (72%) resulted in fruit dropping that caused substantial injury and bruise. Generally, anthracnose caused by C. gloeosporioides of avocado fruit was prevalent in producer orchards that aggravated by traditional harvest and postharvest practices coupled to inadequate transportation and storage facilities at wholesaler and retailer shops with subsequent decay and loss of avocado fruits.     

Bionomics of Anopheles spp. (Diptera: Culicidae) in a malaria endemic region of Sukabumi,West Java,Indonesia

A 15‐month bionomic study of Anopheles species was conducted in two ecologically distinct villages (coastal and upland) of Sukabumi District, West Java, Indonesia from June 2006 to September 2007. Mosquitoes were captured using human‐landing collections at both sites. During the study, a total of 17,100 Anopheles mosquitoes comprising 13 Anopheles species were caught: 9,151 at the coastal site and 7,949 at the upland site. Anopheles barbirostris, Anopheles maculatus, and Anopheles vagus were the predominant species caught at the coastal site, and Anopheles aconitus, Anopheles barbirostris, and An. maculatus predominated in the upland site. Overall, species were exophagic at both sites, but there was variation between species. Anopheles aconitus was endophagic at the coastal site, exophagic at the upland site, collected most often in April 2007 and had a peak landing time between 22:00 and 23:00. Anopheles sundaicus was only collected at the coastal site, exophagic, collected most often in October 2006, and had a peak landing time between 19:00 and 20:00. Potential malaria vector species such An. aconitus, An. maculatus, and An. sundaicus were present throughout the year. None of the 7,770 Anopheles tested using CSP‐ELISA were positive for malaria, although the risk for malaria outbreaks in Sukabumi district remains high.     

Larvicidal efficacy of Aloe barbadensis and Cannabis sativa against the malaria vector Anopheles stephensi (Diptera: Culicidae)

Anopheles stephensi is the primary vector of malaria, an endemic disease in India. An effort to control An. stephensi larvae by leaf extracts of Aloe barbadensis (Liliaceae) and Cannabis sativa (Moraceae) was made under laboratory conditions. A carbon tetrachloride extract of A. barbadensis was the most effective of all the extracts tested for larvicidal activity against the anopheline larvae, with LC₅₀ 15.58 and 8.04 p.p.m. after 24 and 48 h of exposure, respectively. Thus, the leaf extract of A. barbadensis has active components that could be useful as a larvicide of ecocongenial nature against malaria vectors.     

Role of species composition in malaria transmission by the Anopheles funestus group (Diptera: Culicidae) in Ghana

Malaria remains a public health problem in Ghana, with Anopheles gambiae and Anopheles funestus as the predominant vectors. While much information exists on the species composition of An. gambiae, very little exists for An. funestus. This study was carried out to determine the species composition of An. funestus Giles populations from three ecological areas in Ghana and investigate their role in malaria transmission. Mosquitoes were collected using human landing and pyrethrum spray methods. A total of 10,254 Anopheles individuals were collected, out of which An. funestus constituted 53.6% (5,496). An. funestus sensu stricto (s.s.) and Anopheles lessoni were identified as the only members of the An. funestus group in all three ecological areas. All 62 sporozoite positive specimens that were identified as An. funestus s.s. were highly anthropophilic with a human blood index in the range of 80–96%, whereas more than 83% of the An. leesoni had fed on either bovine, goat, or sheep. Malaria transmission was higher in the Sahel savannah area than the rest of the ecological zones, with An. funestus s.s. being implicated as a vector of malaria in all ecological zones. Anopheles leesoni occurred in all the ecological areas but played no role in malaria transmission. The study established the importance of An. funestus s.s. in malaria transmission in Ghana.     

Pyrethroid-treated bednet effects on mosquitoes of the Anopheles gambiae complex in The Gambia

The response of Anopheles gambiae complex mosquitoes to men sleeping under insecticide-impregnated or untreated bednets in six verandah trap huts was studied during the dry season in The Gambia. With this type of hut it was possible to collect live and dead indoor-resting mosquitoes and estimate the number of wild mosquitoes which entered, bloodfed on man, and exited each night. Bednets were treated with emulsions targetted to leave deposits of 25 mg/m2 lambda-cyhalothrin, or 5, 50 or 500 mg/m2 permethrin, diluted from emulsifiable concentrates (EC), or a blank formulation similar to the EC except that the permethrin was omitted; the sixth net was left untreated. Nets and sleepers were rotated between huts on different nights, the design being based on a series of Latin squares and conducted double-blind. Permethrin-impregnated bednets deterred mosquitoes from entering the huts. The degree of deterrency was proportional to the dosage of permethrin. This effect was also caused by the blank formulation and therefore attributed to other components of the formulation, rather than to the permethrin itself. The net impregnated with 500 mg permethrin per square metre gave the best individual protection, reducing mosquito bloodfeeding by 91% compared with untreated nets. However, lambda-cyhalothrin was proportionately more insecticidal than permethrin at doses of equivalent deterrency. At this stage of research, it remains conjectural whether chemical deterrency or killing of malaria vectors is better for community protection.     

印度疟疾媒介库态按蚊卵黄蛋白原基因的克隆与生物信息学分析（英文）

卵黄蛋白原（vitellogenin, Vg)是主要的卵黄蛋白前体, 在雌虫血餐之后在脂肪体内大量合成。卵黄蛋白原的调节元件已经被用于驱动蚊子（与寄生虫发生最大相互作用的场所）中抗寄生基因的组织特异性表达。不过, 迄今为止, 对在印度引起60%~70%疟疾发生的库态按蚊Anopheles culicifacies中的内源启动子尚未进行过分析。本研究通过PCR扩增了包括5′端上游调节区在内的库态按蚊A. culicifacies卵黄蛋白原基因, 并命名为AncuVg (GenBank登录号为JN113091)。它含有一个大约6.2 kb的开放阅读框, 编码2 052个氨基酸, 具有一个16个氨基酸残基的推断的信号肽。也含有一个N_Vitellogenin区和一个VWF型D区, 这两个区在其他昆虫卵黄蛋白原中也保守。估计多肽分子量为238.0 kDa, 含有4个共有的(RXXR/S)切割位点, C端附近有一个GL/ICG基序, 其后是9个半胱氨酸残基和1个位于GL/ICCG基序上游第18个氨基酸残基处的DGXR 基序。在推断的氨基酸序列上发现3个聚丝氨酸区, 其中2个位于氨基端, 1个位于羧基端。根据同义密码子相对使用概率值, 通过有效密码子数, 测定了蚊子卵黄蛋白原基因密码子的偏倚性程度。也预测了库态按蚊A. culicifacies Vg的三维结构。分析了AncuVg基因, 以理解Vg基因的转录调节。对Vg基因5′端上游区进行的系统发育分析表明, 它们聚类于蚊子的3大分枝。也用各种生物信息学工具分析分析了Vg的同源性和特征。     

Laboratory Evaluation of the Lethal Efficacy of an Insect Growth Regulator, Pyriproxyfen against Anopheles sinensis (Diptera, Culicidae) Larvae

The granular formulation of 0.5% pyriproxyfen is a mosquito insect growth regulator larvicide. It was evaluated for residual activity for killing and/or emergence inhibition against 3rd stage larvae of Anopheles sinensis in the laboratory. Mortality rates ranged from 60.9% to 79.7% at 0.01, 0.05 and 0.1 mg/L of water during the first seven days. Except for the lowest concentration (0.01 mg/L) mortality rates were greater than 80% from day 8 through 28. Mortality rates for 0.01 mg/L concentrations were 70.4% from days 8‐14, and thereafter exceeded 80% through day 28. The residual activity gradually increased post‐treatment since the granular formulation of pyriproxyfen has a gradual solubility and increases concentration of active ingredient over time. The mortality rates of An. sinensis larvae and pupae eventually reached 100% for the three (0.01, 0.05, 0.1 mg/L) concentrations.     

Isolation and characterization of microsatellite loci in the mosquito Anopheles stephensi Liston (Diptera: Culicidae)

The mosquito Anopheles stephensi is an important malaria vector in India, Pakistan, Iran and Afghanistan. Differences in egg morphology and chromosomal characters have been described between urban and rural forms of this mosquito but the population genetic structure remains unclear. In India this species is mainly urban, rural populations are largely zoophilic and not thought to transmit malaria. In eastern Afghanistan and the Punjab and Northwest Frontier Province, Pakistan, it is the major malaria vector. We have developed primers for 16 microsatellite loci to assist in defining the population structure and epidemiological importance of this mosquito.     

Identification of candidate volatiles that affect the behavioural response of the malaria mosquito Anopheles gambiae sensu stricto to an active kairomone blend: laboratory and semi‐field assays

Anopheles gambiae Giles sensu stricto (Diptera: Culicidae) is the most important vector of human malaria in sub‐Saharan Africa, affecting the lives of millions of people. Existing tools such as insecticide‐treated nets and indoor‐residual sprays are not only effective, but also have limitations as a result of the development of resistance to insecticides and behavioural adaptations in biting time. Therefore, novel mosquito‐control tools are needed. Odour‐releasing traps have a potential for both trapping and surveillance purposes. Based on the outcome of ex vivo gene expression assays and in vivo electrophysiological assays on hundreds of volatile organic compounds, 29 ‘candidate behaviourally‐disruptive organic compounds' (cBDOCs) are selected, belonging to 10 chemical categories, to be assayed in the laboratory using dual‐choice olfactometers for the ability to modify the ‘attractiveness’ (i.e. significantly more insects being caught in the associated trap) of a basic volatile blend consisting of ammonia, lactic acid and tetradecanoic acid but without additional carbon dioxide. Compounds that either reduce or increase trap catches by the basic blend in the olfactometer experiments are tested under African conditions in choice experiments in a semi‐field facility in Kenya. The release rates of cBDOCs are determined gravimetrically to allow the calculation of aerial concentrations at the trap outlet. Aerial concentrations in the sub‐p.p.m. range are reported for the first time to influence mosquito behaviour. The results of the olfactometer and semi‐field assays generally correspond. Under semi‐field conditions, three compounds, 3‐heptanol (0.025 p.p.m.), 2‐methylpropanal (0.05 p.p.m.) and 4,5‐dimethylthiazole (0.73 p.p.m.), significantly increase trap catches relative to the basic blend consisting of ammonia, lactic acid, tetradecanoic acid and carbon dioxide. 2‐Acetylthiophene, 2‐nonanone and 2‐phenylethanol decrease the number of mosquitoes caught at all concentrations tested under semi‐field conditions. These compounds hold promise as attractants and spatial repellents to be applied in mosquito control programmes.     

土荆芥的石油醚提取物及精油对冈比亚按蚊的杀虫活性

在室内条件下测试了土荆芥Chenopodium ambrosioides 叶片的石油醚提取物及精油对冈比亚按蚊Anopheles gambiae 0～2 d龄成虫和1～4龄幼虫的毒性。每一龄期各取30头幼虫分别接触50,100,250,500,750和1 000 mg/L提取物和精油,于24 h和48 h后记录死亡幼虫的数目。30头成虫在密封的长方形玻璃笼子中分别接触0.8,1.6,2.4 μL/L土荆芥精油蒸气, 24 h后记录死亡率。结果表明,测试的提取物和精油对冈比亚按蚊的各龄幼虫和成虫均具有毒性。48 h致死中浓度（LC50）的测试结果表明,石油醚提取物对1龄幼虫的毒性最强（14.89 mg/L）,其次是对4龄幼虫（18.90 mg/L）,对3龄幼虫的毒性最低（183.77 mg/L）; 精油对4龄幼虫毒性最强（36.62 mg/L）,其次是对1龄幼虫（90.75 mg/L）。推算的土荆芥精油对冈比亚按蚊的LC50为1.01 μL/L。本研究揭示了土荆芥对冈比亚按蚊的防治潜力。     

Effects of larval density and feeding rates on larval life history traits in Anopheles gambiae s.s. (Diptera: Culicidae)

Mosquito larval habitat determines the fitness, survivorship, fecundity, and vector capacity of emerging adults. We manipulated larval density and food provisioning in the laboratory to determine their effects on a number of life history parameters of Anopheles gambiae. Larval mortality was positively correlated with larval density. In addition, increased density skewed sex ratios that favored females.     

Feeding and survival of the malaria vector Anopheles gambiae on plants growing in Kenya

The propensity of the malaria vector mosquito Anopheles gambiae Giles (Diptera: Culicidae) to ingest sugars from various plants, and subsequent survival rates, were assessed with laboratory-reared males and females offered eight species of plants commonly cultivated and/or growing wild in western Kenya. In cages (no-choice bioassay), mosquitoes given the opportunity to feed on castorbean (Ricinus communis L.) had the longest survival times (mean and median survival time of 6.99 +/- 0.23 and 5.67 +/- 0.17 days, respectively), comparable to mosquitoes given 6% glucose (mean and median survival time of 8.70 +/- 0.23 and 6.67 +/- 0.33 days, respectively). Survival rates of An. gambiae were low on the other plants, comparable to mosquitoes given only water. Three plants: sweet potato (Ipomoea batatas L.), wild sage (Lantana camara L.) and castorbean provided levels of sugar ingestion by both sexes of An. gambiae detectable using the cold anthrone method, showing a positive correlation between median survival and sugar consumption (Spearman rank correlation coefficient = 0.905, P < 0.0001). Equal numbers of males and females were released in an enclosed semi-field screenhouse system containing a range of local plants, but no host for blood, and allowed to feed ad libitum: 6.7 +/- 0.5% (11/64) of those recaptured were found to contain detectable fructose (all females). Common plants are clearly a viable source of nutrition for adult female An. gambiae, as well as males, and may constitute and important resource for this important malaria vector.