A Role for the Insulin Receptor in the Suppression of Dengue Virus and Zika Virus in Wolbachia-Infected Mosquito Cells

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Evaluating perceptions of risk in mosquito experts and identifying undocumented pathways for the introduction of invasive mosquito species into Europe

In several reported cases of the entry of invasive mosquito species (IMSs) into Europe, the introduction was associated with a specific pathway of introduction or dispersal. The identification of potential pathways for the introduction of IMSs and evaluations of the importance of the different pathways are key to designing proper surveillance strategies to promptly detect and control introductions in non‐infested areas. The main goals of the present study were to identify other, previously undocumented, pathways of introduction into Europe, and to identify mosquito experts' perceptions regarding control measures against IMS introductions via different documented pathways. At the European Mosquito Control Association (EMCA) conference in Montenegro in March 2017, a questionnaire was distributed among meeting participants to collect expert data. Results showed that ground transportation (by cars, trucks, etc.), passive natural dispersal and the shipping of used tyres are perceived as the most likely pathways. Introduction via aircraft did not appear to be well known and was not perceived as probable. This study shows that there were no pathways unknown to European experts that could lead to cryptic introductions into the experts' countries. Furthermore, the findings demonstrated that the perceived efficacy of surveillance and control is key to overcoming the constraints experienced and to supporting the implementation of actions against introductions.     

Physiological Recordings and RNA Sequencing of the Gustatory Appendages of the Yellow-fever Mosquito Aedes aegypti

Electrophysiological recording of action potentials from sensory neurons of mosquitoes provides investigators a glimpse into the chemical perception of these disease vectors. We have recently identified a bitter sensing neuron in the labellum of female Aedes aegypti that responds to DEET and other repellents, as well as bitter quinine, through direct electrophysiological investigation. These gustatory receptor neuron responses prompted our sequencing of total mRNA from both male and female labella and tarsi samples to elucidate the putative chemoreception genes expressed in these contact chemoreception tissues. Samples of tarsi were divided into pro-, meso- and metathoracic subtypes for both sexes. We then validated our dataset by conducting qRT-PCR on the same tissue samples and used statistical methods to compare results between the two methods. Studies addressing molecular function may now target specific genes to determine those involved in repellent perception by mosquitoes. These receptor pathways may be used to screen novel repellents towards disruption of host-seeking behavior to curb the spread of harmful viruses.     

Human sperm injection into the perivitelline space (SI-PVS) of hamster oocytes: Effect of sperm pretreatment by calcium-ionophore A23187 and freezing-thawing on the penetration rate and polyspermy

Calcium-ionophore A23187 and freezing-thawing were used as sperm treatments before human sperm injection into the perivitelline space (SI-PVS) of hamster oocytes. The penetration rate (PR) was higher when SI-PVS was performed with calcium-ionophore-treated (28%) or frozen-thawed (51%) sperm than with untreated sperm (8%). Optimal PR occurred when five calcium-ionophore-treated (57%) or frozen-thawed (71%) sperm were injected under the zona pellucida. When the sperm:egg ratio was 1:1, PR was higher for calcium-ionophore-treated (18.5%) or frozen-thawed (27.8%) sperm than for untreated sperm (0.0%). Calcium-ionophore sperm treatment had no effect on the polyspermic oocyte rate (POR) or the mean number of swollen sperm nuclei per penetrated oocyte (Pd) or per injected sperm (SR). This may result from premature oocyte activation induced by Ca-ionophore. However, POR was higher with frozen-thawed (74%) than with untreated (50%) or Ca-ionophore-treated (50%) sperm. Whatever the sperm treatment, there was a trend toward a lower SR as the number of injected sperm increased. Cytoplasmic regulation of polyspermy in the hamster oocyte is discussed.     

《遗传资源获取与惠益分享的名古屋议定书》核心内容解读及其生效预测

《生物多样性公约》缔约方大会第十次会议通过的《生物多样性公约关于遗传资源获取和公平公正地分享由遗传资源利用产生惠益的名古屋议定书》是实现《生物多样性公约》确保公平公正的分享因利用遗传资源而产生的惠益目标的里程碑式文件,将从多个方面对国际社会产生影响。对议定书进行解读、分析和评估对于决定是否签署和批准议定书至关重要。根据分析,议定书核心内容分为遵守、公平公正的惠益分享、遗传资源的获取、术语、范围、特殊考虑和与遗传资源相关的传统知识等7个方面。本文对这些核心内容进行了解读,分析了其影响,预测了议定书生效的可能性,并提出了应对措施。     

学科前沿知识贯穿于微生物学教学过程的思考与实践——以反硝化作用教学为例

根据好氧反硝化作用机理研究进展和好氧反硝化细菌在污水脱氮处理工艺中的应用,结合微生物生态的教学内容,在提出教材重点教学内容与学科前沿知识优化整合方案的基础上,进一步阐述了在不增加课堂理论教学时数的前提下,如何通过学生课外自主学习环节进行好氧反硝化作用教学的做法,发现课外专题学习与课堂教学紧密结合是提高微生物学教学效果的有效方法,也是创新型应用型人才培养的有效途径。     

Rates of DNA Duplication and Mitochondrial DNA Insertion in the Human Genome

The hundreds of mitochondrial pseudogenes in the human nuclear genome sequence (numts) constitute an excellent system for studying and dating DNA duplications and insertions. These pseudogenes are associated with many complete mitochondrial genome sequences and through those with a good fossil record. By comparing individual numts with primate and other mammalian mitochondrial genome sequences, we estimate that these numts arose continuously over the last 58 million years. Our pairwise comparisons between numts suggest that most human numts arose from different mitochondrial insertion events and not by DNA duplication within the nuclear genome. The nuclear genome appears to accumulate mtDNA insertions at a rate high enough to predict within-population polymorphism for the presence/absence of many recent mtDNA insertions. Pairwise analysis of numts and their flanking DNA produces an estimate for the DNA duplication rate in humans of 2.2 × 10^–9 per numt per year. Thus, a nucleotide site is about as likely to be involved in a duplication event as it is to change by point substitution. This estimate of the rate of DNA duplication of noncoding DNA is based on sequences that are not in duplication hotspots, and is close to the rate reported for functional genes in other species.     

Effects of global warming on mosquitoes & mosquito‐borne diseases and the new strategies for mosquito control

Global warming has shortened mosquitoes’ lifecycle period and increased the disease transmission rates by mosquito vectors. We reviewed only three mosquito‐borne diseases: malaria, dengue fever, and the Japanese encephalitis. Billions of people get infected with those diseases and millions of people die every year. Although we struggle to find the most effective way to control mosquitoes using various methods (including pesticides), mosquito‐borne diseases are still among the most serious problems being faced. This paper, therefore, reviews the strategies for controlling mosquitoes. The use of pesticides to control mosquitoes might have more negative effects on humans and environments than benefits. Although the development of genetically modified (GM) mosquitoes raises new hopes for effective mosquito control, it will take longer to assess the risks to humans and environments. Furthermore, there has been concern about the possible adverse effects from the release of GM mosquitoes into the environment. The various mosquito traps may not be as effective at controlling only female mosquito populations. Therefore, new strategies for the control of mosquitoes are vital. The smart mosquito counter device was developed by Korean Centre for Disease Control (KCDC) in 2013. The mosquito pest control office is able to set up the appropriate mosquito control strategies by using quantitative mosquito information. The smart device will bring mosquito control in line with modern smart generation technology and the device will also soon be able to identify different mosquito species. This new strategy will change the methods of mosquito control and will provide beneficial effects toward sustainable nature and human health.     

A technique and apparatus for placement of test substances into the intestines of mosquito larvae

A method for injection of substances into the intestinal lumen via the anal pore is described for fourth instar larvae of Aedes aegypti. Both oral and anal injections were possible with fourth instar larvae of Toxorhynchites amboinensis.

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Résumé On a pu introduire à partir du pore anal des échantillons liquides d'environ 1/3 l dans la lumière intestinale de larves du 4e stade d'Aedes aegypti. Un colorant injecté s'est réparti dans l'intestin postérieur et l'intestin moyen.11 l de solution ont pu être injectés oralement ou analement dans des larves de 4e stade de Toxorhynchites amboinensis. Les mortalités moyennes ont été de 20% (0 à 30%) pour des larves de A. eegypti et de 4% (0 à 10%) pour les larves de T. amboinensis. Nous avons évalué l'interêt de cette technique pour l'étude de l'action directe de toxines et d'autres substances sur les cellules intestinales. Nous avons souligné les possiblités de prélèvement local et de mesures physiologiques (pH, potentiel électrique transépithélial).

     

Study of the Insects Associated with the Floodwater Mosquito Ochlerotatus Albifasciatus (Diptera: Culicidae) and their Possible Predators in Buenos Aires Province,Argentina

Insects associated with the floodwater mosquito Ochlerotatus albifasciatus(Diptera: Culicidae) were studied from intermittent puddles in temperate Argentina in an attempt to detect the main predators. Forty-one taxa occurred in the puddles from spring to fall. Coleoptera and Diptera were dominant and diverse. Ephemeroptera and Odonata were scarce in numbers and species, and Heteroptera occurred in low numbers of species and high abundance of individuals. The main predators of immature O. albifasciatus were detected on the basis of relative abundance (ISA index), ecological dominant groups, and species association (I index). Liodessus sp. and Rhantus signatus signatus(Coleoptera: Dytiscidae) were the most abundant predators in the puddles and Liodessus sp., Lancetes marginatus(Dytiscidae) and Psorophora ciliata(Culicidae) were the most frequent. Liodessus sp. and O. albifasciatus were the best associated species in all seasons. Liodessus sp. was indistinctly associated with all larval instars and pupae, while R. signatus signatus, L. marginatus and Psorophora ciliata were more associated with the pupal stage of the mosquito. Desmopachria concolor(Coleoptera: Dytiscidae), Tropisternus lateralis limbatus and Tropisternus setiger(Coleoptera: Hydrophilidae) were other potential predators inhabiting the puddles, but their relevance was minor.     

Physicochemical and Safety Evaluation of 5-Aminolevulinic Acid in Novel Liposomes as Carrier for Skin Delivery

In this study we successfully entrapped 5-aminolevulinic acid (ALA) in liposome, although it exists as a zwitter ion. A molar ratio of 2:1:2.5 phosphatidyle-thanolamine (PE)/cholesterol/sodium stearate represented the best condition to achieve high entrapment efficiency (29.37 ± 1.21%), and the average vehicle size was 133.6 ± 2.8 nm. After 32 days of storage, the vehicle sizes of formulations with PE series were still approximately less than 200 nm. The safety of liposomes was tested and ensured both with regard to cellular cytotoxicity and erythrocyte hemolysis. Safety studies showed that liposome formulations did not affect cell viability except when both potassium stearate and sodium oleate were added. Moreover, PE and PE/cholesterol did not damage human erythrocytes in this study. The range of the hemolytic effect caused by liposomes was 5 to 37% and the effect was dependent on the amount of sodium stearate added to the formulation. According to the release rates and skin penetration of ALA liposomes in vitro, PE/cholesterol/sodium stearate liposomes might increase skin penetration, and it was shown that penetration across the stratum–corneum (sc) layer was the rate-limiting process. Images from confocal laser scanning microscopy (CLSM) confirmed the great potency of liposomes for delivering ALA into skin.     

Sperm penetration in vitro into ovarian and tubal oocytes from mice of the inbred KE and C57 strains

The method of in vitro fertilization was applied to test a previous suggestion that the lowered fertilizability of the tubal oocytes of female KE strain mice and the high resistance of their zona pellucida to proteolytic enzymes, are due to the premature cortical reaction taking place near the time of ovulation. Therefore higher fertilizability of ovarian oocytes is expected. The effectiveness of F₁ hybrid sperm penetration into ovarian and tubal KE oocytes confirmed these assumptions. The ovarian KE oocytes recovered 9–10 hours after administration of human chorionic gonadotropin (HCG) showed significantly higher penetrability (70–83%) than did the tubal oocytes recovered 12 hours after HCG (about 50%) and 14–16 hours after HCG (20%). Similar results were obtained with C57 oocytes. Sperm penetration into ovarian oocytes (10 hours after HCG) was much more effective (67%) than into tubal oocytes (18%); this finding correlated with more rapid zona dissolution by chymotrypsin. On the basis of these results one might speculate that premature cortical reaction takes place also in the C57 strain.     

Human skin microbiota and their volatiles as odour baits for the malaria mosquito Anopheles gambiae s.s

Host seeking by the malaria mosquito Anopheles gambiae Giles sensu stricto (Diptera: Culicidae) is mainly guided by volatile chemicals present in human odours. The skin microbiota plays an important role in the production of these volatiles, and skin bacteria grown on agar plates attract An. gambiae s.s. in the laboratory. In this study, the attractiveness of volatiles produced by human skin bacteria to An. gambiae s.s. was tested in laboratory, semi‐field, and field experiments to assess these effects in increasing environmental complexity. A synthetic blend of 10 compounds identified in the headspace of skin bacteria was also tested for its attractiveness. Carbon dioxide significantly increased mosquito catches of traps baited with microbial volatiles in the semi‐field experiments and was therefore added to the field traps. Traps baited with skin bacteria caught significantly more An. gambiae s.s. than control traps, both in the laboratory and semi‐field experiments. Traps baited with the synthetic blend caught more mosquitoes than control traps in the laboratory experiments, but not in the semi‐field experiments. Although bacterial volatiles increased mosquito catches in the field study, trapping several mosquito vector species, these effects were not significant for An. gambiae s.l. It is concluded that volatiles from skin bacteria affect mosquito behaviour under laboratory and semi‐field conditions and, after fine tuning, have the potential to be developed as odour baits for mosquitoes.     

Non-endocytic penetration of core histones into petunia protoplasts and cultured cells: a novel mechanism for the introduction of macromolecules into plant cells

The results of the present work demonstrate that core histones are able to penetrate the plasma membrane of plant cells. Confocal microscopy has revealed that incubation of petunia protoplasts with fluorescently labeled core histones resulted in cell penetration and nuclear import of the externally added histones. Intracellular accumulation was also confirmed by an ELISA-based quantitative method using biotin-labeled histones. Penetration into petunia protoplasts and cultured cells was found to be non-saturable, occurred at room temperature and at 4 °C and was not inhibited by Nocodazole. Furthermore, penetration of the biotinylated histone was neither blocked by the addition of an excess of free biotin molecules, nor by non-biotinylated histone molecules. All these results clearly indicate that the observed uptake is due to direct translocation through the cell plasma membrane and does not occur via endocytosis. Our results also show that the histones H2A and H4 were able to mediate penetration of covalently attached BSA molecules demonstrating the potential of the histones as carriers for the delivery of macromolecules into plant cells. To the best of our knowledge, the findings of the present paper demonstrate, for the first time, the activity of cell penetrating proteins (CPPs) in plant cells.     

人表皮角质形成细胞自噬功能细胞模型的构建及鉴定

目的: 建立稳定表达GFP-LC3的人永生化角质形成细胞HaCaT细胞系。方法: 将构建的pcDNA3.1-GFP-LC3 真核表达载体转入HaCaT细胞,经G418筛选稳定表达的细胞系。HaCaT细胞中GFP-LC3的表达分别用荧光显微镜与Western blot方法检测,并利用该稳定表达的细胞系观察验证Rapamycin对细胞发生自噬透射电镜超微结构的变化。结果: 获得了3株转染并经G418反复筛选的HaCaT细胞系,在倒置荧光显微镜下观察可见绿色荧光细胞的表达率在95% 以上,Western blot结果证实了GFP-LC3融合蛋白的表达。Western blot和激光共聚焦显微镜均证明Rapamycin可以诱导自噬的发生。透射电镜细胞超微结构的观察表明Rapamycin可以有效地诱导HaCaT-LC3细胞自噬的发生。结论: 成功构建GFP-LC3稳定表达的HaCaT系,该细胞系可以作为研究人角质形成细胞自噬功能的一种细胞模型。     

Inherited DNA Repair Defects Disrupt the Structure and Function of Human Skin

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