海南岛按蚊的鉴定和迷走按蚊的进化研究

目的 阐明海南岛某些按蚊的分类地位,探讨迷走按蚊与其近缘种的亲缘关系. 方法 对采自海南岛8个地点的按蚊依据形态和分子特征进行鉴别研究,应用的分子特征包括核糖体DNA(ribosomal DNA,rDNA)第2内转录间隔区(second internal transcribed spacer,ITS2)和28S第3编码区(third domain,D3)序列,综合分析迷走按蚊及其近缘种的ITS2和D3序列,构建系统发育树. 结果 本研究共鉴定按蚊407只,其中84.9％的成蚊分子鉴定与依据形态鉴定的结果一致.在228条迷走按蚊的ITS2序列中,部分个体在2个位点存在单碱基套峰(G/A),出现的频率分别为57.9％、64.8％,同时有两个套峰的占样本数的57.0％,其他位点保守.迷走按蚊的D3序列,在种内无碱基差异.结合本研究和文献的迷走按蚊及其近缘种ITS2序列构建的ML树,显示迷走按蚊与浅色按蚊MC/D亲缘关系近,与另一支的浅色按蚊B、圣代克按蚊复合体的亲缘关系较远.关于未订名种NBC (Anophelessp.NBG)的分类地位,依据分子特征鉴定其为浅色按蚊B,可能是圣代克按蚊复合体新的成员种.结论 鉴定形态在个体间变异大的种类时,客观的分子特征更具重要性,迷走按蚊与浅色按蚊MC/D具较近的亲缘关系.     

In vivo transposition of Minos, a Drosophila mobile element, in mammalian tissues

Transposable elements have been used widely in the past 20 years for gene transfer and insertional mutagenesis in Drosophila. Transposon-based technology for gene manipulation and genomic analysis currently is being adopted for vertebrates. We tested the ability of Minos, a DNA transposon from Drosophila hydei, to transpose in mouse tissues. Two transgenic mouse lines were crossed, one expressing Minos transposase in lymphocytes under the control of the CD2 promoter/locus control region and another carrying a nonautonomous Minos transposon. Only mice containing both transgenes show excision of the transposon and transposition into new chromosomal sites in thymus and spleen cells. In addition, expression of Minos transposase in embryonic fibroblast cell lines derived from a transposon-carrying transgenic mouse resulted in excision of the transposon. These results are a first step toward a reversible insertional mutagenesis system in the mouse, opening the way to develop powerful technologies for functional genomic analysis in mammals.     

Dusk to dawn activity patterns of anopheline mosquitoes in West Timor and Java, Indonesia

Malaria is a serious health issue in Indonesia. We investigated the dusk to dawn anopheline mosquito activity patterns, host-seeking and resting locations in coastal plain, hilly and highland areas in West Timor and Java. Adult mosquitoes were captured landing on humans or resting in houses or animal barns. Data analyzed were: mosquito night-time activities; period of peak activity; night-time activity in specific periods of time and for mosquito resting locations. Eleven species were recorded; data were sparse for some species therefore detailed analyses were performed for four species only. In Java Anopheles vagus was common, with a bimodal pattern of high activity. In West Timor, its activity peaked around midnight. Other species with peak activity around the middle of the night were An. barbirostris and An. subpictus. Most species showed no biting and resting preference for indoors or outdoors, although An. barbirostris preferred indoors in West Timor, but outdoors in Java. An. aconitus and An. annularis preferred resting in human dwellings; An. subpictus and An. vagus preferred resting in animal barns. An. barbirostris preferred resting in human dwellings in West Timor and in animal barns in Java. The information is useful for planning the mosquito control aspect of malaria management. For example, where mosquito species have peak activity at night indoors, bednets and indoor residual spraying should reduce malaria risk, but where mosquitoes are most active outdoors, other options may be more effective.     

Larvicidal activity of Bacillus sphaericus 2362 against Anopheles nuneztovari, Anopheles darlingi and Anopheles braziliensis (Diptera, Culicidae).

In this present study, preliminary data was obtained regarding the mortality rate of the Amazonian anophelines, Anopheles nuneztovari, Anopheles darlingi and Anopheles braziliensis when subjected to treatment with Bacillus sphaericus strain 2362, the WHO standard strain. Initially, experiments were conducted to test the mortality rate of the three species of anopheline larvae. The third larval instar of An. nuneztovari and the second and third larval instars of An. darlingi proved to be the least susceptible. In other experiments, the same three mosquito species were tested with the standard strain 2362, An. nuneztovari was the least susceptible to this insect pathogen, while An. braziliensis was the most susceptible. This latter species showed a difference in the level of LC50 concentration, when compared to the former, of 2.4, 2.5 and 1.8 in readings taken 24, 48 and 72 hours after exposure to the bacillus.     

中华按蚊吸血趋性和吸血前后活动情况的观察

目的观察中华按蚊吸血趋性和吸血前后活动情况,为改进灭蚊措施,提高防制效果提供依据。方法利用室外人、牛饵帐诱捕法进行中华按蚊吸血趋性观察,采用实验小屋法观察蚊虫吸血前后活动情况。结果室内人饵帐捕获中华按蚊约是室外捕蚊数的2.99倍,差异有统计学意义(χ2=15.12,P<0.01);室内外牛饵帐诱捕蚊数相近,差异无统计学意义(χ2=0.69,P>0.05)。牛房傍晚3h的捕蚊数为白天栖息该室内蚊虫数的12.59倍,在室内吸血前后停息1~10min者约占50%,停息30min以内者占80%以上,差异均有统计学意义(P<0.05);停息60min以上者仅占10%,差异无统计学意义(P>0.05)。结论中华按蚊具有偏外栖性,这是滞留喷洒杀灭效果不理想的重要原因。基于其吸血前后室内短暂停留的行为习性,应选用无驱避性且具有熏杀作用的速效高效杀虫剂杀灭室内中华按蚊。     

Gonadal activity and chromosomal constitution of in vitro generated embryos

Chromosomal analysis of pre-implantation embryos was carried out in patients with a poor prognosis of full term pregnancy, which underwent induction of multiple follicular growth. In all, 1034 embryos generated from 191 stimulated cycles were screened for nine chromosome aneuploidy by using the multicolour fluorescence in situ hybridisation technique. Thirty-five percent of the diagnosed embryos were chromosomally normal, whereas the remaining presented with numerical abnormalities, which made them not suitable for transfer. The results obtained confirmed that the incidence of abnormalities is mostly dependent on age; however, monosomy and trisomy are more frequent in poor responders. Accordingly, the pregnancy rate per started cycle was significantly higher in women with a normal response to gonadotropic stimulation (33% vs. 8%, P<0. 001). These findings indicate that poor responder patients are physiologically exposed not only to reduced chances of implantation, but also to an increased risk of spontaneous abortion and trisomic pregnancies.     

Increased bone resorption in moderate smokers with low body weight: the Minos study

Tobacco was found to be a risk factor for osteoporosis, mainly in postmenopausal women. We studied the effect of smoking on bone mineral density (BMD) and bone turnover in a cohort of 719 men, aged 51-85 yr, composed of 83 current smokers, 405 former smokers, and 231 men who never smoked. Most current and former smokers were moderate smokers (median, 10 cigarettes/d). Current smokers were younger, thinner, and drank more coffee and more alcoholic beverages. After adjustment for age, body weight, alcohol intake, and caffeine intake, current and former smokers had similar BMD, except at the forearm. Former smokers had lower BMD compared with never-smokers at most skeletal sites. Men who had smoked more than 7120 packs (third quartile) had lower BMD of total hip (P < 0.01) and distal forearm (P = 0.03) compared with men in the 2 lower tertiles. In the 3 groups, levels of bone formation markers did not differ. After adjustment for confounding variables, levels of urinary markers of bone resorption (beta-isomerized C-terminal telopeptide, free and total deoxypyridinoline) were higher in the current smokers than in former smokers and never-smokers. Concentrations of T, total 17beta-E2, and androstenedione were higher, whereas that of 25-hydroxyvitamin D was lower, in current smokers. When men were divided according to tertiles of body weight, increased bone resorption, decreased BMD and biochemical indexes of secondary hyperparathyroidism were observed in current smokers in the lowest tertile of body weight (<75 kg) compared with the never-smokers, but not in men in the two highest tertiles of body weight. Current smokers had a higher prevalence of vertebral deformities after adjustment for age and body weight (13% vs. 5%; P < 0.005). In summary, in moderate smokers with low body weight (<75 kg), increased bone resorption, not matched by increased bone formation, results in decreased BMD and an increased prevalence of vertebral deformities. In this group, low serum 25-hydroxyvitamin D and secondary hyperparathyroidism may explain, at least partly, the effect of tobacco on bone turnover. In former smokers, bone resorption is not increased, but BMD remains lower compared with that in never-smokers.     

Physiology of desiccation resistance in Anopheles gambiae and Anopheles arabiensis

Desiccation resistance and water balance were studied in the adult female mosquitoes Anopheles arabiensis and Anopheles gambiae sensu stricto. When the two species were reared from egg to adult under identical conditions, An. arabiensis had significantly higher desiccation resistance than did An. gambiae. Data are presented that indicate that this difference in desiccation resistance is associated with a higher body water content prior to desiccation in An. arabiensis. No differences in rate of water loss during desiccation or water content at death were observed. Measurements of metabolic rate and respiratory pattern also showed no statistically significant differences between the species. This study provides the first physiologic measurements of desiccation resistance in adults of these species and offers insights into the physiologic differences associated with differential resistance to desiccation stress.     

Neutrophil-activating activity and platelet-activating factor synthesis in cytokine-stimulated endothelial cells: reduced activity in growth-arrested cells

The reactivity of endothelial cells (ECs) to proinflammatory cytokines is critically important for the pathogenesis of vascular diseases. Here, we studied functional alterations of human ECs during culture under a confluent condition; i.e., the alterations of neutrophil-activating activity, platelet-activating factor (PAF) synthesis, and granulocyte-macrophage colony-stimulating factor (GM-CSF) production in cytokine-stimulated ECs. Human umbilical vein-derived ECs exhibited the increased activity in neutrophil activation, PAF synthesis, and GM-CSF production when stimulated by proinflammatory cytokines such as interleukin-1 beta (IL-1 beta) and tumor necrosis factor-alpha (TNF-alpha). The activity of cytokine-stimulated ECs to stimulate superoxide release in human neutrophils and to produce PAF declined markedly in parallel as ECs became growth-arrested during culture under a confluent condition. By contrast, GM-CSF production induced by cytokine stimulation was modestly increased, and up-regulation of intercellular adhesion molecule-1 (ICAM-1) and activation of mitogen-activated protein kinases were not altered. The neutrophil-activating activity of cytokine-stimulated ECs was dependent on PAF synthesis and GM-CSF production from ECs. These findings indicate that the reduced neutrophil-activating activity in growth-arrested ECs may be, at least in part, ascribed to down-regulation of PAF synthesis.     

JunB inhibits proliferation and transformation in B-lymphoid cells

The activator protein 1 (AP-1) member JunB has recently been implicated in leukemogenesis. Here we surveyed human lymphoma samples for expression of JunB and other AP-1 members (c-Jun, c-Fos, Fra1, JunD). JunB was strongly expressed in T-cell lymphomas, but non-Hodgkin B-cell lymphomas do not or only weakly express JunB. We therefore asked whether JunB acted as a negative regulator of B-cell development, proliferation, and transformation. We used transgenic mice that expressed JunB under the control of the ubiquitin C promoter; these displayed increased JunB levels in both B- and T-lymphoid cells. JunB transgenic cells of B-lymphoid, but not of T-lymphoid, origin responded poorly to mitogenic stimuli. Furthermore, JunB transgenic cells were found to be less susceptible to the transforming potential of the Abelson oncogene in vitro. In addition, overexpression of JunB partially protected transgenic mice against the oncogenic challenge in vivo. However, transformed B cells eventually escaped from the inhibitory effect of JunB: the proliferative response was similar in explanted tumor-derived cells from transgenic animals and those from wild-type controls. Our results identify JunB as a novel regulator of B-cell proliferation and transformation.     

Water quality and breeding habitats of anopheline mosquito in northwestern Thailand

Malaria transmission is dependent upon many hydrology-driven ecological factors that directly affect the vectorial competence, including the presence of suitable habitats for the development of anopheline larvae. Larval habitats were identified and characterized at three malaria endemic villages (Ban Khun Huay, Ban Pa Dae, and Ban Tham Seau) in Mae Sot district, Tak Province, in northwestern Thailand between July 2002 and June 2003. The Global Positioning System (GPS) was used to provide precise locational data for the spatial distribution of anopheline mosquito larvae and their habitats. Ten habitat categories were identified. Eighteen adult Anopheles species were identified from larvae in all the surveyed habitats. An. minimus was the most common species throughout the year. The relationship between eight abiotic variables (temperature, hardness, carbon dioxide, dissolved oxygen, nitrate, phosphate, silica and pH) and the abundance of four major species of malaria vectors (An. (Cel.) dirus, An. (Cel.) minimus, An. (Cel.) maculatus, and An. (Cel.) sawadwongporni), and six species of non-vectors (An. (Cel.) kochi, An. (Cel.) jamesii, An. (Ano.) peditaeniatus, An. (Ano.) barbirostris, An. (Ano.) campestris, and An (Cel.) vagus) larvae was investigated. The results from the multiple regression models suggest that hardness, water temperature and carbon dioxide are the best predictor variables associated with the abundance of An. minimus larvae (p < 0.001); water pH for An. dirus larvae (p < 0.001); temperature and pH for An. kochi larvae (p < 0.01); temperature and silica concentration for An. jamesii larvae (p < 0.001); dissolved oxygen and silica concentration for An. campestris larvae (p < 0.001); and pH and silica concentration for An. vagus larvae (p < 0.001). We could not identify key environmental variables for An. maculatus, An. sawadwongporni, An. peditaeniatus, and An. barbirostris.     

Larvicidal and pupicidal activity of spinosad against the malarial vector Anopheles stephensi

ObjectiveTo investigate the larvicidal and pupicidal activity of spinosad against Anopheles stephensi Listen.MethodsSpinosad from the actinomycete, Saccharopolyspora spinosa was tested against Anopheles stephensi at different concentrations (0.01, 0.02, 0.04, 0.06 and 0.08 ppm.), and against first to fourth instar larvae and pupae.ResultsThe larval mortality ranged from 36.1±1.7 in (0.01 ppm) to 79.3±1.8 (0.08 ppm) the first instar larva. The LC₅₀ and LC₉₀ values of first, second, third and fourth instar larva were 0.001, 0.031, 0.034, 0.036 and 0.0113, 0.102, 0.111, 0.113, respectively. The pupal mortality ranged from 33.0±2.0 (0.01 ppm) to 80.0±0.9 (0.08 ppm). The LC₅₀ and LC₉₀ values were 0.028 and 0.1020, respectively. The reduction percentage of Anopheles larvae was 82.7%, 91.4% and 96.0% after 24, 48, 72 hours, respectively, while more than 80% reduction was observed after 3 weeks.ConclusionsIn the present study spinosad effectively caused mortality of mosquito larvae in both the laboratory and field trial. It is predicted that spinosad is likely to be an effective larvicide for treatment of mosquito breeding sites.     

拜虫杀对中华按蚊和嗜人按蚊毒杀效果的实验观察

目的观察拜虫杀对中华按蚊和嗜人按蚊的毒杀效果。方法采用拜虫杀10、15、20mg/m2(有效成分)浸泡棉纱布和尼龙纱布,以实验室饲养的中华按蚊和嗜人按蚊为试虫,进行击倒力、致死力和持效观察。结果采用拜虫杀10、15、20mg/m2浸泡两种材料蚊帐,中华按蚊和嗜人按蚊接触药帐后的半数击倒时间(KT50)在3.98~7.30min之间,被击倒的蚊虫恢复饲养24h,死亡率均为100%;接触药帐3min,恢复饲养24h,蚊虫死亡率均>90%;杀虫剂浸泡过的蚊帐悬挂180d后,对媒介按蚊的击倒力仍在8.57~16.31min之间,且恢复饲养24h,死亡率仍为100%。结论拜虫杀对中华按蚊和嗜人按蚊有很强的杀灭效果,持效可达180d以上。     

In vivo leukemic transformation: cytogenetic evidence of in vivo leukemic transformation of engrafted marrow cells.

A 4-year-old girl with acute myeloblastic leukemia was treated with 50 mg/kg cyclophosphamide daily for 4 days before being given 1.7 X 10(8) bone marrow cells/kg from her HL-A identical, MLC nonreactive, cytogenetically normal brother. The patient died 92 weeks after the marrow graft. Postmortem examination showed increased myeloblasts and promyelocytes. Cytogenetic studies before transplantation showed that all sex determination metaphases had an XX pattern, and 41% of the hyperdiploid metaphases had an additional 19-20(F) chromosome. At autopsy all hyperdiploid metaphases with XY pattern and 43% of the hyperdiploid metaphases with an XX pattern had an additional F chromosome. Occasional metaphases with 47, XX, + F or 47, XY, + F were seen during the follow-up studies. These findings indicated that an acute leukemia had developed in the XY cell line of this artifically induced sex chimeric child. This suggests that a leukemic stimulus other than that proposed to be induced by total-body irradiation existed in this patient and transformed the engrafted cells.     

Developmental expression of vitamin K-dependent gamma-carboxylase activity in zebrafish embryos: effect of warfarin

Vitamin K-dependent gamma-carboxylation is an essential posttranslational modification required for the functional activity of coagulation proteins such as factors VII, IX, X, and prothrombin. Warfarin, an inhibitor of vitamin K-dependent gamma-carboxylation, was used in earlier work on adult zebrafish to provide evidence for the presence of vitamin K-dependent carboxylase in zebrafish. Here we demonstrate the presence of vitamin K-dependent carboxylase activity in zebrafish by directly assaying the microsomal fraction prepared from adult, unfertilized eggs, and embryos from different developmental stages. Gamma-carboxylase activity was detected both before and after fertilization of embryos and the activity levels remained relatively constant from 6 h postfertilization (hpf) through other advanced stages of development. The expression of activity in the early embryos (0-6 hpf) may be due to the presence of maternal protein since the activity was detected even in the unfertilized eggs. Gamma-carboxylase activity in the eggs as well as early embryos suggested that vitamin K-dependent carboxylase is important throughout development. The detection of vitamin K-dependent carboxylase mRNA by RT-PCR and inhibitor studies using warfarin confirmed these activity results. Further, these studies provide a basis for selecting warfarin-resistant zebrafish mutants in order to find genes regulating gamma-carboxylase activity including the yet unidentified vitamin K-epoxide reductase.     

Community sleeping pattern and anopheline biting in southeastern iran: a country earmarked for malaria elimination

Abstract. An important variable in determining the vectorial capacity of malaria mosquito species is the degree of mosquito-human contact. This parameter can be affected by community sleeping behavior and the host-feeding habits of vectors. A cross-sectional study of 775 randomly selected inhabitants, including 385 Baluchi residents and 390 Afghani refugees, was conducted in a malarious area in Sabaz District, Sistan-Baluchestan Province, southeastern Iran. In addition, monitoring of human landing periodicity of main malaria vectors was carried out during an entire transmission season. Afghanis and Baluchis showed diversity in sleeping behavior. Most (79.6%) respondents were familiar with symptoms of malaria and also aware of an association between mosquitoes and malaria. Despite this familiarity, 94.6% of Afghan refugees, 74.8% of Baluch residents, and 87.2% of study participants did not use self-protection preventive measures. Overall, only 8.8% of participants reported using bed nets regularly. Surveyed persons used bed nets mainly during second quarter of night. Three major species of malaria vectors (Anopheles culicifacies, An. fluviatilis, and An. stephensi) started biting by sunset and continued throughout the night. The results of present study indicated that synchronization of encounters between inhabitants and mosquito vectors was caused by poor self-protection and sleeping behavior of inhabitants. In addition, diversity in culture and behavior of the two communities may cause the prevalence of malaria to be different between them. Therefore, promoting awareness of self-protection against mosquito bites could promote community participation in malaria elimination program in this malaria-endemic region.     

Increased melanizing activity in Anopheles gambiae does not affect development of Plasmodium falciparum

Serpins are central to the modulation of various innate immune responses in insects and are suspected to influence the outcome of malaria parasite infection in mosquito vectors. Three Anopheles gambiae serpins (SRPN1, -2, and -3) were tested for their ability to inhibit the prophenoloxidase cascade, a key regulatory process in the melanization response. Recombinant SRPN1 and -2 can bind and inhibit a heterologous phenoloxidase-activating protease and inhibit phenoloxidase activation in vitro. Using a reverse genetics approach, we studied the effect of SRPN2 on melanization in An. gambiae adult females in vivo. Depletion of SRPN2 from the mosquito hemolymph increases melanin deposition on foreign surfaces such as negatively charged Sephadex beads. As reported, the knockdown of SRPN2 adversely affects the ability of the rodent malaria parasite Plasmodium berghei to invade the midgut epithelium and develop into oocysts. Importantly, we tested whether the absence of SRPN2 from the hemolymph influences Plasmodium falciparum development. RNAi silencing of SRPN2 in an An. gambiae strain originally established from local populations in Yaoundé, Cameroon, did not influence the development of autochthonous field isolates of P. falciparum. This study suggests immune evasion strategies of the human malaria parasite and emphasizes the need to study mosquito innate immune responses toward the pathogens they transmit in natural vector-parasite combinations.