Interference between bunyaviruses in Aedes triseriatus mosquitoes

Inhibition of the replication of alternate California serogroup bunyaviruses in Aedes triseriatus mosquitoes has been observed for mosquitoes previously infected with La Crosse (LAC) virus. By contrast, prior infection of mosquitoes with LAC virus did not interfere significantly with the subsequent infection and replication of Guaroa bunyavirus (Bunyamwera serogroup), or heterologous viruses such as West Nile flavivirus, or vesicular stomatitis rhabdovirus.     

Transduction of Aedes aegypti mosquitoes with vectors derived from Aedes densovirus

Aedes densovirus (AeDNV)-based constructs that express green fluorescent protein (GFP) from either the P7 or the P61 promoter were made. The construct in which GFP protein was expressed as a fusion protein to the C-terminus of NS1 (NS1-GFP) showed the highest level of GFP expression. This hybrid NS1-GFP protein preserved the biological functions of the parental proteins: it showed GFP fluorescence, it stimulated expression from the virus promoters, and it facilitated rescue and replication of the cloned AeDNV genome. Similar to NS1, the hybrid NS1-GFP localized in the nucleus predominantly in a punctate pattern. Transducing virus particles carrying the NS1-GFP gene infected mosquito larvae. Expression of GFP was detected as early as 48 h postinfection and in larval and pupal stages. Midgut, hindgut, and Malpighian tubule cells expressed GFP soon after transduction. However, the anal papillae were the most commonly infected organ system. The anal papillae are syncytia and regulate ion concentration in the hemolymph of mosquito larvae, and they might be a novel route of mosquito larvae infection with densoviruses.     

Dirofilaria repens microfilariae in Aedes vexans mosquitoes in Slovakia

In this study, we screened field-caught mosquitoes for presence of Dirofilaria spp. by using a polymerase chain reaction (PCR) assay. Potential occurrence of Dirofilaria repens and Dirofilaria immitis microfilariae was examined in 3,600 mosquitoes of eight species (Aedes vexans, Aedes cinereus, Aedes rossicus, Culex pipiens, Culiseta annulata, Ochlerotatus sticticus, Ochlerotatus cantans and Ochlerotatus caspius) collected from five locations in two districts (Kosice and Trebisov) of Eastern Slovakia, endemic region of canine dirofilariasis. Collection of mosquitoes was performed between May and August 2012 in premises known to be inhabited by Dirofilaria-infected dogs. PCR assays were performed on 72 pools, each pool containing 50 mosquitoes of the same species, collected on the same location. Each pool was examined separately for the presence of D. immitis and D. repens, respectively. A positive finding of D. repens was recorded in one pool of A. vexans mosquitoes collected in Ko?ické Ol?any village. Minimum infection rate in A. vexans was 1:1,750, i.e. 0.57 per 1,000 mosquitoes. The identity of D. repens was confirmed by direct sequencing of PCR product which has shown 100 % homology with sequence attributed to D. repens (GenBank accession number AJ271614). This study represents the first molecular evidence of D. repens microfilariae in mosquitoes in Slovakia and highlights a need for better surveillance of zoonotic dirofilariasis in central Europe.     

Reassortment of La Crosse and Tahyna bunyaviruses in Aedes triseriatus mosquitoes

Experiments were conducted to determine if La Crosse (LAC) and Tahyna (TAH) viruses reassort in Aedes triseriatus mosquitoes and to determine the genotypic frequencies of viruses selected by in vivo vector interactions. A molecular hybridization technique was used to analyze progeny viruses. Probes specific for the La Crosse L, M and S segments (pLAC4.16: LAC L RNA; pLAC4.27: LAC M RNA; pLAC4C-26: LAC S RNA) were used to determine the parental origin of the progeny RNA segments. Following infection with a mixture of LAC and TAH viruses, mosquitoes were held for 23 days extrinsic incubation, then assayed for reassortment. Individual progeny viruses were isolated by plaque assay and propagated in BHK-21 cells. Cytoplasmic RNA was extracted from the cells, blotted in triplicate to Nytran, and each blot was hybridized with 32P-labelled pLAC4.16, pLAC4.27 or pLAC4C-26 to determine the parental origin of each RNA segment. High frequency reassortment occurred in these mosquitoes. All of the expected genotypes resulting from a cross of LAC and TAH were obtained from these mosquitoes. Genotypic frequencies of 708 virus isolates from 39 mosquitoes were: LLL, 150 (21%); LLT, 71 (10%); LTL, 39 (5.5%); LTT, 109 (15%); TTT, 259 (36%); TTL, 16 (2.2%); TLT, 55 (7.8%); TLL, 9 (1.2%).     

The sensilla of Aedes and Anopheles mosquitoes and their importance in repellency

The aim of this study was to detect the role of some mosquito organs in their sensation of repellent materials. A total of 250 females (15 days old) of the target species Aedes aegypti and Anopheles stephensi were prepared and divided into five groups: group 1, without antenna; group 2, without maxillary bulbs; group 3, without proboscis; group 4, without frontal tarsus; and group 5, normal females as control. A mixture of five oils containing Litsea cubeba 1%, Melaleuca leucadendron 1%, Melaleuca quinquenervia 1%, Viola odorata 1%, and Nepeta cataria 1% was included in a complex solvent containing 20% genapol, 10% polyethylene glycol, 20% ethanol, and 50% water. Furthermore, Bayrepel was used in this experiment at a 20% concentration in the same solvent. Pure water was used as control in this study. The test was carried out by spreading 100 μl of the repellent material or water on a 30-cm² exposure area of a human volunteer’s arm. In A. aegypti, the biting and landing percentages increased significantly in those mosquito groups that lacked some organs (especially maxillary bulbs), while in A. stephensi, it became not clear which organ is responsible for perception of repellents.     

Classical fowl plague virus reproduction in the body of Aedes aegypti mosquitoes

The results of the studies on fowl plague virus (FPV, Rostok strain) reproduction in Aedes aegypti mosquitoes are presented. The virus-containing allantoic fluid was inoculated intrathoracally in volumes of 0.1 and 0.2 microliter. The virus was isolated in chick embryos and could be detected at 5--14 days after inoculation. After inoculation of 0.1 microliter of virus it could be detected in doses of 0.5, 2.0, 1.75 Ig2 ID50, after inoculation of 0.2 microliter--in doses of 5, 1.5, and 0.5 Ig2 ID50.     

Detection of chikungunya virus antigen in Aedes albopictus mosquitoes by enzyme-linked immunosorbent assay

Double-antibody sandwich and modified sandwich systems of enzyme-linked immunosorbent assay for detecting chikungunya virus antigen present in female mosquitoes, Aedes albopictus (Oahu strain), were evaluated as simple and rapid methods of selection of a highly susceptible mosquito line. Both assays were capable of detecting 3.9 X 10(1) ng (4.0 X 10(6) PFU) or more of the purified antigen. An inhibition system was less sensitive, and a direct system with adsorption of test specimens on the solid phase was not useful. Positive reactions were observed in 16 (48.5%) of 33 infected mosquitoes with with 10(6) to 10(7) PFU, which correspond to the highly susceptible group of this strain. Mosquitoes with less than 10(6) PFU were all negative, indicating the usefulness of the sandwich techniques for identifying high-titered mosquitoes.     

Genetic differentiation of invasive Aedes albopictus by RAPD-PCR: implications for effective vector control

Aedes albopictus is an invasive mosquito species of great concern to public health as it is responsible for the biological transmission of several pathogens causing dengue, chikungunya, yellow fever, etc. In 2009, this mosquito was detected for the first time in Agra City. This study represents the first genetic analysis of A. albopictus from India. Random amplified polymorphic DNA (RAPD) was used to study the genetic structure of A. albopictus in four populations from different larval habitats. Seven RAPD primers produced 141 loci. The results displayed rich genetic variation among larval populations which is evident from high value of genetic differentiation (G _ST), i.e. 0.280, indicating a very great genetic differentiation. Effective migration rates were observed to be 1.28, depicting a limited gene flow. According to analysis of molecular variance (AMOVA), the genetic distance between populations was significant (P?<?0.05), showing a very high intrapopulation variation (96 %) with only 4 % variation among populations. Average genetic distances between populations were also calculated using PopGene software. Nei’s average genetic distance between these populations was 0.112 (0.05–0.18). The cluster analysis technique of unweighted pair-group mean analysis (UPGMA) method of arithmetic averages was used to develop the phylogenetic tree which clearly shows two clusters of different larval habitats. The findings highlight high genetic differentiation indicating a slight migration rate confirming the recent introduction of this species in Agra region.     

Integrated control of peridomestic larval habitats of Aedes and Culex mosquitoes (Diptera: Culicidae) in atoll villages of French Polynesia

An integrated larval mosquito control program was carried out in Tiputa village on Rangiroa atoll of French Polynesia. Mosquito abundance before and after treatment was compared with the abundance in an untreated village. Mosquito larval habitats consisted of large concrete or polyurethane cisterns, wells, and 200-liter drums. Depending on the target species, larval habitat category, its configuration, and purpose (drinking consumption or not), abatement methods consisted of sealing the larval habitats with mosquito gauze, treating them with 1% Temephos, covering the water with a 10-cm thick layer of polystyrene beads or introducing fish (Poecillia reticulata Rosen & Bailey). All premises of the chosen village were treated and a health education program explained basic mosquito ecology and the methods of control. A community health agent was trained to continue the control program at the end of the experiment. Entomological indices from human bait collections and larval surveys indicated that mosquito populations were reduced significantly, compared with concurrent samples from the untreated control village, and that mosquito control remained effective for 6 mo after treatment. Effects of the treatment were noticed by the inhabitants in terms of a reduction in the number of mosquito bites. In the Polynesian context, such control programs may succeed in the long-term only if strong political decisions are taken at the village level, if a community member is designated as being responsible for maintaining the program, and if the inhabitants are motivated sufficiently by the mosquito nuisance to intervene.     

Isolation of the causative agent of Karelian fever from Aedes sp. mosquitoes

The results of isolation of an Alphavirus strain from Aedes sp. mosquitoes collected in the focus of Karelian fever are presented. According to the results of serological studies, the isolated strain LEIV-9298 Karelia belongs to the antigenic complex Sindbis-Western equine encephalomyelitis. The appurtenance of the isolated agent to the Alphavirus genus has been confirmed by electron microscopic examinations. A rise of antibody titres to LEIV-9298 Karelia virus in paired sera of subjects with a history of Karelian fever allows it to be considered the causative agent of this disease.     

Vector competence of Aedes aegypti and Aedes albopictus (Diptera: Culicidae) for dengue virus in the Florida Keys

In 2009-2011, Monroe County in southern Florida experienced locally acquired and traveler-imported focal dengue outbreaks. Aedes aegypti (L.) is the primary vector of dengue virus (DENV) worldwide, is prevalent in Monroe County, and is the suspected vector in Florida. Ae. albopictus (Skuse) is also known to be an important vector of DENV and this species is ubiquitous in Florida; however, it is not yet established in Monroe County. Florida Ae. aegypti (Key West and Stock Island geographic colonies) and Ae. albopictus (Vero Beach geographic colony) were fed blood containing 3.7 Log10 plaque-forming unit equivalents of DENV serotype 1 isolated from a patient involved in the Key West, FL, outbreak in 2010. Mosquitoes were maintained at extrinsic incubation temperatures of 28 or 30 degrees C for an incubation period of 14 d. Vector competence was assessed using rates of infection (percent with virus-positive bodies), dissemination (percent infected with virus-positive legs), and transmission (percent infected with virus-positive saliva). No significant differences were observed in rates of infection or dissemination between Ae. aegypti or Ae. albopictus at either extrinsic incubation temperature. Transmission was observed only at 28 degrees C in both Ae. aegypti (Key West) and Ae. albopictus. The assessment of local mosquito populations for their DENV vector competence is essential and will aid mosquito control operators interested in pinpointing specific vector populations for control. The extent to which vector competence is affected by seasonal changes in temperature is discussed and provides baseline risk assessment data to mosquito control agencies.     

Mechanical transmission of Bacillus anthracis by stable flies (Stomoxys calcitrans) and mosquitoes (Aedes aegypti and Aedes taeniorhynchus).

We evaluated the potential of stable flies, Stomoxys calcitrans, and two species of mosquitoes, Aedes aegypti and Aedes taeniorhynchus, to transmit Bacillus anthracis Vollum 1B mechanically. After probing on Hartley guinea pigs with a bacteremia of ca. 10(8.6) CFU of B. anthracis per ml of blood, individual or pools of two to four stable flies or mosquitoes were allowed to continue feeding on either uninfected guinea pigs or A/J mice. All three insect species transmitted lethal anthrax infections to both guinea pigs and mice. Both stable flies and mosquitoes transmitted anthrax, even when they were held at room temperature for 4 h after exposure to the bacteremic guinea pig before being allowed to continue feeding on the susceptible animals. This study confirms that blood-feeding insects can mechanically transmit anthrax and supports recent anecdotal reports of fly-bite-associated cutaneous human anthrax. The potential for flies to mechanically transmit anthrax suggests that fly control should be considered as part of a program for control of epizootic anthrax.     

Repellency effect of forty-one essential oils against Aedes, Anopheles, and Culex mosquitoes

Since ancient times, plant products were used in various aspects. However, their use against pests decreased when chemical products became developed. Recently, concerns increased with respect to public health and environmental security requiring detection of natural products that may be used against insect pests. In this study, 41 plant extracts and 11 oil mixtures were evaluated against the yellow fever mosquito, Aedes aegypti (Linnaeus), the malaria vector, Anopheles stephensi (Liston), and the filariasis and encephalitis vector, Culex quinquefasciatus (Say) (Diptera: Culicidae) using the skin of human volunteers to find out the protection time and repellency. The five most effective oils were those of Litsea (Litsea cubeba), Cajeput (Melaleuca leucadendron), Niaouli (Melaleuca quinquenervia), Violet (Viola odorata), and Catnip (Nepeta cataria), which induced a protection time of 8 h at the maximum and a 100% repellency against all three species. This effect needs, however, a peculiar formulation to fix them on the human skin.