Detection of Chikungunya virus in Aedes aegypti during 2011 outbreak in Al Hodayda, Yemen

In October 2010, the Ministry of Public Health and Population reported an outbreak of dengue-like acute febrile illness in Al Hodayda governorate. By January 2011, a total of 1542 cases had been recorded from 19 of the 26 districts in the governorate with 104 purportedly associated deaths. In response this event, in January 2011 entomological investigations aimed at identifying the primary vector and the epidemic associated etiological agent were carried out. Based on the reported cases and the progress of the outbreak in the governorate, mosquito collection was undertaken in two of the most recent outbreak areas; Al Khokha district (130 km south of Al Hodayda) and Al Muneera district (100 km north). Mosquito adults were collected from houses using BG-sentinel? traps, aspiration of resting mosquitoes and knock-down spraying. Indoor and outdoor containers adjacent to the houses were inspected for larvae. Subsequently mosquito pools were analyzed by RT-PCR for detection of the four dengue virus serotypes (DENV-1, DENV-2, DENV-3, DENV-4), and for Chikungunya virus (CHIKV). Aedes aegypti was the dominant mosquito species collected. Four pools represent 40% of the tested pools, all containing adult female Ae. aegypti, were positive for CHIKV. Three CHIKV isolates were obtained from the RNA positive mosquito pools and identified by rRT-PCR. This finding marks the first record of CHIKV isolated from Ae. aegypti in Yemen. The larval container and Breteau indices in the visited localities surveyed were estimated at 53.8 and 100, respectively. The emergence of this unprecedented CHIKV epidemic in Al Hodayda is adding up another arboviral burden to the already existing vector-borne diseases. Considering the governorate as one focal port in the Red Sea region, the spread of the disease to other areas in Yemen and in neighboring countries is anticipated. Public health education and simple measures to detect and prevent mosquito breeding in water storage containers could prevent and reduce the spread of mosquito-borne viruses like CHIKV and DENV in Yemen.     

Dengue Knowledge and Preventive Practices in Iquitos,Peru

As part of a cluster-randomized trial to evaluate insecticide-treated curtains for dengue prevention in Iquitos, Peru, we surveyed 1,333 study participants to examine knowledge and reported practices associated with dengue and its prevention. Entomological data from 1,133 of these households were linked to the survey. Most participants knew that dengue was transmitted by mosquito bite (85.6%), but only few (18.6%) knew that dengue vectors bite during daytime. Most commonly recognized dengue symptoms were fever (86.6%), headache (76.4%), and muscle/joint pain (67.9%). Most commonly reported correct practices for mosquito control were cleaning homes (61.6%), using insecticide sprays (23%), and avoiding having standing water at home (12.3%). Higher education was associated with higher knowledge about dengue, including transmission and vector control. Higher socioeconomic status was associated with increased reported use of preventive practices requiring money expenditure. We were less likely to find Aedes aegypti eggs, larvae, or pupae in households that had < 5-year-old children at home. Although dengue has been transmitted in Iquitos since the 1990s and the Regional Health Authority routinely fumigates households, treats domestic water containers with larvicide, and issues health education messages through mass media, knowledge of dengue transmission and household practices for prevention could be improved.     

Naturally-Acquired Dengue Virus Infections Do Not Reduce Short-Term Survival of Infected Aedes aegypti from Ho Chi Minh City,Vietnam

Transmission of dengue virus (DENV) from mosquito to human is dependent upon the survival of the mosquito beyond the virus extrinsic incubation period. Previous studies report conflicting results of the effects of DENV on Aedes aegypti survival. Here, we describe the effect of DENV on the short-term survival (up to 12 d) of 4,321 Ae. aegypti mosquitoes blood-fed on 150 NS1-positive dengue patients hospitalized in the Hospital for Tropical Diseases, Ho Chi Minh City, Vietnam. Mosquito survival was not different between cohorts that fed upon blood from which 0% of mosquitoes became DENV infected (N = 88 feeds), or 100% became infected (N = 116 feeds). Subgroup analysis also did not reveal serotype-dependent differences in survival, nor a relationship between survival and human plasma viremia levels. These results suggest that DENV infection adds minimal cost to Ae. aegypti, an important finding when parameterizing the vector competence of this mosquito.     

Stable transformation of the yellow fever mosquito, Aedes aegypti, with the Hermes element from the housefly

The mosquito Aedes aegypti is the world’s most important vector of yellow fever and dengue viruses. Work is currently in progress to control the transmission of these viruses by genetically altering the capacity of wild Ae. aegypti populations to support virus replication. The germ-line transformation system reported here constitutes a major advance toward the implementation of this control strategy. A modified Hermes transposon carrying a 4.7-kb fragment of genomic DNA that includes a wild-type allele of the Drosophila melanogaster cinnabar (cn) gene was used to transform a white-eyed recipient strain of Ae. aegypti. Microinjection of preblastoderm mosquito embryos with this construct resulted in 50% of the emergent G₀ adults showing some color in their eyes. Three transformed families were recovered, each resulting from an independent insertion event of the cn⁺-carrying transposon. The cn⁺ gene functioned as a semidominant transgene and segregated in Mendelian ratios. Hermes shows great promise as a vector for efficient, heritable, and stable transformation of this important mosquito vector species.     

Effects of extract of soapnut Sapindus emarginatus on esterases and phosphatases of the vector mosquito, Aedes aegypti (Diptera: Culicidae)

Our earlier investigations with kernels from the soapnut Sapindus emarginatus revealed it as a new source of botanical biocide with potent antimosquito activity, as evident from the proven unique ability of the aqueous kernel extract to kill all the developmental stages of three important vector mosquito species, Aedes aegypti, Anopheles stephensi and Culex quinquefasciatus. This extract was also found to be safe for two non-target aquatic insects. As a sequel to these findings, we have further examined quantitative and qualitative changes in total proteins, esterases, and phosphatases in whole body homogenates of fourth instar larvae and pupae of A. aegypti exposed to this extract at an appropriate threshold time for its lethal effect to gain insights into the impact of the botanical biocide on biochemical characteristics of the target vector mosquito at two distinct developmental stages. The profiles of proteins, esterases (acetylcholinesterse, α- and β-carboxylesterases), and phosphatases (acid and alkaline) exhibited distinct patterns of variation during normal development of fourth instar larvae and pupae, indicating intrinsic difference in biochemical features between these two developmental stages of A. aegypti. Upon exposure of the larvae to the extract, significant reduction in the activities of acetylcholinesterse, β-carboxylesterase, and acid phosphatases were recorded, whereas the total proteins, α-carboxylesterase and alkaline phosphatase activities were unaffected. By contrast, only alkaline phosphatase activity was significantly affected in pupae exposed to the extract. Analysis of these enzymes in native PAGE revealed that they exist in isoforms in both the larvae and pupae. The alterations in the levels of enzymatic activities observed from the quantitative assays of various enzymes were reflected by the respective zymograms with perceptible differences in the intensity and the number of bands detected especially with β-carboxylesterase, acid and alkaline phosphatase activity between the control and exposed test organisms. Despite the fact that the soapnut kernel extract causes mortality of both the larvae and pupae of A. aegypti, the findings of this study demonstrate that the impact of this extract is most pronounced in various enzyme profiles of the larvae rather than the pupae. Such discrepancy implicates the presence of unique biochemical mechanisms in the pupae of mosquito for detoxification of botanical biocides.     

Immuno-suppressive effects of aqueous extract of soapnut Sapindus emarginatus on the larvae and pupae of vector mosquito,Aedes aegypti

We recently reported the presence of potent anti-mosquito activity in aqueous kernel extract of the soapnut, Sapindus emarginatus, and demonstrated its impact on marker enzymes in larvae and pupae of the vector mosquito, Aedes aegypti. As a sequel to these findings, the present study elucidates immunotoxicity of this extract with respect to hemocyte-mediated cellular immune responses in fourth instar larvae and pupae as well as cuticular melanization reaction in the larvae of A. aegypti. The exposure of these two developmental stages of the mosquito to the soapnut extract at a lethal threshold concentration neither affected hemocyte viability tested up to 3 h in vitro nor did it influence the hemocyte count. By contrast, exposure of the mosquito larvae and pupae to this extract significantly reduced the ability of their hemocytes to bind yeast cells, an important early event in the process of non-self recognition by immune cells. Consequently, the phagocytic activity of these hemocytes against yeast cells was also found to be adversely affected upon exposure of larvae and pupae to the extract. Besides, a perceptible initial delay in melanization reaction at the injured site of the cuticle in the extract-exposed larvae was observed. All these findings demonstrate, for the first time, the immuno-suppressive potential of a botanical biocide in the vector mosquito.     

Effect of Dengue-2 Virus Infection on Protein Expression in the Salivary Glands of Aedes aegypti Mosquitoes

Dengue virus (DENV) is the most important mosquito-transmitted flavivirus that is transmitted throughout the tropical and subtropical regions of the world. The primary mosquito vector of DENV in urban locations is Aedes aegypti. Key to understanding the transmission of DENV is the relationship between pathogen and vector. Accordingly, we report our preliminary characterization of the differentially expressed proteins from Ae. aegypti mosquitoes after DENV infection. We investigated the virus–vector interaction through changes in the proteome of the salivary glands of mosquitoes with disseminated DENV serotype 2 (DENV-2) infections using two-dimensional gel electrophoresis and identification by mass spectrometry. Our findings indicate that DENV-2 infection in the Ae. aegypti salivary gland alters the expression of structural, secreted, and metabolic proteins. These changes in the salivary gland proteome highlight the virally influenced environment caused by a DENV-2 infection and warrant additional investigation to determine if these differences extend to the expectorated saliva.     

Adulticidal properties of Pithecellobium dulce (Roxb.) Benth. (Family: Fabaceae) against dengue vector,Aedes aegypti (Linn.) (Diptera: Culicidae)

To examine the toxicity of mosquito adulticidal activity of different solvent leaf and seed extracts of Pithecellobium dulce (P. dulce) against dengue vector, Aedes aegypti (Ae. aegypti).MethodsAdulticidal efficacy of the crude leaf and seed extracts of P. dulce with five different solvents like benzene, hexane, ethyl acetate, methanol and chloroform was tested against the five to six day old adult female mosquitoes of Ae. aegypti. The adult mortality was observed after 24 h under the laboratory conditions.ResultsAmong the tested solvents the maximum efficacy was observed in the leaf and seed methanol extract. The LC₅₀ and LC₉₀ values of P. dulce leaf and seed extract against adults of Ae. aegypti were 218.64, 257.99 mg/L and 426.05, 507.73 mg/L, respectively. No mortality was observed in controls. The Chi-square values were significant at P<0.05 level.ConclusionsFrom the results it can be concluded the crude extract of P. dulce leaf and seed was an excellent potential for controlling dengue vector mosquito, Ae. aegypti.     

Hot temperatures can force delayed mosquito outbreaks via sequential changes in Aedes aegypti demographic parameters in autocorrelated environments

Aedes aegypti L. (Diptera: Culicidae) is a common pantropical urban mosquito, vector of dengue, Yellow Fever and chikungunya viruses. Studies have shown Ae. aegypti abundance to be associated with environmental fluctuations, revealing patterns such as the occurrence of delayed mosquito outbreaks, i.e., sudden extraordinary increases in mosquito abundance following transient extreme high temperatures. Here, we use a two-stage (larvae and adults) matrix model to propose a mechanism for environmental signal canalization into demographic parameters of Ae. aegypti that could explain delayed high temperature induced mosquito outbreaks. We performed model simulations using parameters estimated from a weekly time series from Thailand, assuming either independent or autocorrelated environments. For autocorrelated environments, we found that long delays in the association between the onset of “hot” environments and mosquito outbreaks (10 weeks, as observed in Thailand) can be generated when “hot” environments sequentially trigger a larval survival decrease and over-compensatory fecundity increase, which lasts for the whole “hot” period, in conjunction with a larval survival increase followed by a fecundity decrease when the environment returns to “normal”. This result was not observed for independent environments. Finally, we discuss our results implications for prospective entomological research and vector management under changing environments.     

Is routine dengue vector surveillance in central Brazil able to accurately monitor the Aedes aegypti population? Results from a pupal productivity survey

Objective To assess how well the Aedes aegypti infestation rapid survey, Levantamento Rapido de Indice para Aedes aegypti (LIRAa), is able to accurately estimate dengue vector densities and target the most important breeding sites in Goiania, Brazil. Methods In February 2009, a pupal productivity survey was conducted in 2 districts of the city of Goiania, central Brazil. The results were compared to those of LIRAas conducted in the same districts during the months before and after the pupal productivity survey. Results In the pupal productivity survey, 2 024 houses were surveyed and 2 969 water‐holding containers were inspected. Discarded small water containers most frequently contained immature Ae. aegypti. The most pupal‐productive containers were elevated water tanks, roof gutters and water holding roofs. Combined, these three containers accounted for <40% of all positive containers but produced >70% of all pupae. In the two districts where the pupal productivity survey was conducted, the house indices were 5.1 and 4.6 and the Breteau indices were 5.9 and 6.0. In contrast, the two LIRAs conducted in the same two districts resulted in an average house index of 1.5 and Breteau index of 2.5, with discarded items identified as the most frequently infested container type. Conclusion Both the LIRAa and the pupal productivity survey identified discarded items as being most frequently infested with immature stages of Ae. aegypti, but the pupal productivity survey showed that elevated containers produced the greatest proportion of Ae. aegypti pupae (a proxy measure of adult vector density) and that the values of the Stegomyia indices were substantially underestimated by LIRAa. Although both surveys differ considerably in terms of sampling method and manpower, in the case of this study the LIRAa did not accurately identify or target the containers that were the most important to adult mosquito production.     

Ecology of vector mosquitoes in Sri Lanka--suggestions for future mosquito control in rice ecosystems

Mosquito-borne diseases are a major public health threat in Asia. To explore effective mosquito control strategies in rice ecosystems from the ecological point of view, we carried out ecological analyses of vector mosquitoes in Sri Lanka. During the 18-month study period, 14 Anopheles, 11 Culex, 5 Aedes, 2 Mansonia, and 1 Armigeres species were collected, most of which are disease vectors for malaria, filariasis, Japanese encephalitis, or dengue in Sri Lanka and elsewhere in Asia. The density and occurrence of Anopheles and Culex species were the highest in seepage pools and paddy fields, where the majority of niche overlaps between larval mosquito and aquatic insect species were observed. All 7 aquatic insect species, which are larval mosquito predators, overlapped their niche with both Anopheles and Culex larvae. This suggests that conserving these aquatic insect species could be effective in controlling mosquito vectors in the study site. Correlations between several climatic factors and mosquito density were also analyzed, and weather conditions, including higher temperature, lower relative humidity, and higher wind velocity, were found to affect mosquito oviposition, propagation, and survival. These findings deepen our understanding of mosquito ecology and will strengthen future mosquito control strategies in rice ecosystems in Asia.     

Dynamics of midgut microflora and dengue virus impact on life history traits in Aedes aegypti

Significant morbidity and potential mortality following dengue virus infection is a re-emerging global health problem. Due to the limited effectiveness of current disease control methods, mosquito biologists have been searching for new methods of controlling dengue transmission. While much effort has concentrated on determining genetic aspects to vector competence, paratransgenetic approaches could also uncover novel vector control strategies. The interactions of mosquito midgut microflora and pathogens may play significant roles in vector biology. However, little work has been done to see how the microbiome influences the host's fitness and ultimately vector competence. Here we investigated the effects of the midgut microbial environment and dengue infection on several fitness characteristics among three strains of the primary dengue virus vector mosquito Aedes aegypti. This included comparisons of dengue infection rates of females with and without their normal midgut flora. According to our findings, few effects on fitness characteristics were evident following microbial clearance or with dengue virus infection. Adult survivorship significantly varied due to strain and in one strain varied due to antibiotic treatment. Fecundity varied in one strain due to microbial clearance by antibiotics but no variation was observed in fertility due to either treatment. We show here that fitness characteristics of Ae. aegypti vary largely between strains, including varying response to microflora presence or absence, but did not vary in response to dengue virus infection.     

Female-specific flightless phenotype for mosquito control

Dengue and dengue hemorrhagic fever are increasing public health problems with an estimated 50–100 million new infections each year. Aedes aegypti is the major vector of dengue viruses in its range and control of this mosquito would reduce significantly human morbidity and mortality. Present mosquito control methods are not sufficiently effective and new approaches are needed urgently. A “sterile-male-release” strategy based on the release of mosquitoes carrying a conditional dominant lethal gene is an attractive new control methodology. Transgenic strains of Aedes aegypti were engineered to have a repressible female-specific flightless phenotype using either two separate transgenes or a single transgene, based on the use of a female-specific indirect flight muscle promoter from the Aedes aegypti Actin-4 gene. These strains eliminate the need for sterilization by irradiation, permit male-only release (“genetic sexing”), and enable the release of eggs instead of adults. Furthermore, these strains are expected to facilitate area-wide control or elimination of dengue if adopted as part of an integrated pest management strategy.     

Spatial Clustering of Aedes aegypti Related to Breeding Container Characteristics in Coastal Ecuador: Implications for Dengue Control

Mosquito management within households remains central to the control of dengue virus transmission. An important factor in these management decisions is the spatial clustering of Aedes aegypti. We measured spatial clustering of Ae. aegypti in the town of Borbón, Ecuador and assessed what characteristics of breeding containers influenced the clustering. We used logistic regression to assess the spatial extent of that clustering. We found strong evidence for juvenile mosquito clustering within 20 m and for adult mosquito clustering within 10 m, and stronger clustering associations for containers ≥ 40 L than those < 40 L. Aedes aegypti clusters persisted after adjusting for various container characteristics, suggesting that patterns are likely attributable to short dispersal distances rather than shared characteristics of containers in cluster areas. These findings have implications for targeting Ae. aegypti control efforts.     

Sustained,Area-Wide Control of Aedes aegypti Using CDC Autocidal Gravid Ovitraps

We have shown that the Centers for Disease Control and Prevention (CDC) autocidal gravid ovitraps (AGO trap) reduced the Aedes aegypti population and prevented mosquito outbreaks in southern Puerto Rico. After showing treatment efficacy for 1 year, we deployed three traps per home in an area that formerly did not have traps and in a site that served as the intervention area. Two new areas were selected as reference sites to compare the density of Ae. aegypti without traps. We monitored mosquitoes and weather every week in all four sites. The hypotheses were the density of Ae. aegypti in the former reference area converges to the low levels observed in the intervention area, and mosquito density in both areas having control traps is lower than in the new reference areas. Mosquito density in the former reference area decreased 79% and mosquito density in the new reference areas was 88% greater than in the intervention areas.     

Aedes aegypti in Jamaica,West Indies: container productivity profiles to inform control strategies

Objective To describe the Aedes aegypti container profile in the three parishes of Portland, St. Anns and St. Catherine, Jamaica. Method Traditional stegomyia and pupae per person indices. Results A total of 8855 containers were inspected. A. aegypti were breeding in 19.2% of the 4728 containers in Portland, in 6.7% of the 2639 containers in St. Ann, and in 27.2% of the 1488 containers in Tryhall Heights, St. Catherine. Container types differed between Portland (P > 0.02) on one hand and St. Ann and Tryhall Heights, St. Catherine on the other hand: there were with no vases or potted plants with water saucers in St. Ann and St. Catherine. A. aegypti were breeding in more containers in St. Catherine (38%) (38% in wet season and 21% in the dry season) than in Portland (19%) or St. Ann (6%), both of which had more containers but A. aegypti breeding in fewer: 17.7% and 11.2% in the wet and 20.4% and 3.5% in the dry seasons respectively. The daily production of adult mosquitoes in the three study sites was 1.51, 1.29 and 0.66 adult female mosquitoes per person in Portland, St. Ann and St. Catherine during the dry season and 1.12, 0.23 and 1.04 female mosquitoes per person in the wet season respectively. Conclusion All three communities are at risk for dengue outbreaks and vector control should concentrate on reducing the mosquito populations from the most productive containers before a new dengue virus serotype is introduced into Jamaica.     

Low Aedes aegypti Vector Competence for Zika Virus from Viremic Rhesus Macaques

Despite worldwide efforts to understand the transmission dynamics of Zika virus (ZIKV), scanty evaluation has been made on the vector competence of Aedes aegypti fed directly on viremic human and non-human primates (NHPs). We blood-fed Ae. aegypti from two districts in Rio de Janeiro on six ZIKV infected pregnant rhesus macaques at several time points, half of which were treated with Sofosbuvir (SOF). Mosquitoes were analyzed for vector competence after 3, 7 and 14 days of incubation. Although viremia extended up to eight days post monkey inoculation, only mosquitoes fed on the day of the peak of viremia, recorded on day two, became infected. The influence of SOF treatment could not be assessed because the drug was administered just after mosquito feeding on day two. The global infection, dissemination and transmission rates were quite low (4.09%, 1.91% and 0.54%, respectively); no mosquito was infected when viremia was below 1.26 × 10⁵ RNA copies/mL. In conclusion, Ae. aegypti vector competence for ZIKV from macaques is low, likely to be due to low viral load and the short duration of ZIKV viremia in primates suitable for infecting susceptible mosquitoes. If ZIKV infection in human and macaques behaves similarly, transmission of the Zika virus in nature is most strongly affected by vector density.     

Evidence of Limited Polyandry in a Natural Population of Aedes aegypti

The mosquito Aedes aegypti is a vector of yellow fever, dengue, and chikungunya. Control of the insect is crucial to stop the spread of dengue and chikungunya, so it is critically important to understand its mating behavior. Primarily, based on laboratory behavior, it has long been assumed that Ae. aegypti females mate once in their lifetime. However, multiple inseminations have been observed in semi-field and laboratory settings, and in closely related species. Here, we report the first evidence of polyandry in a natural population of Ae. aegypti. Female Ae. aegypti were captured around the New Orleans, LA, metropolitan area. They were offered a blood meal and allowed to lay eggs, which were reared to the third-instar larval stage. A parentage analysis using four microsatellite loci was performed. Out of 48 families, 3 showed evidence of multiple paternity. An expanded analysis of these three families found that one family group included offspring contributed by three fathers, and the other two included offspring from two fathers. This result establishes that polyandry can occur in a small proportion of Ae. aegypti females in a natural setting. This could complicate future genetic control efforts and has implications for sampling for population genetics.     

Toxicity effect of Delonix elata (Yellow Gulmohr) and predatory efficiency of Copepod,Mesocyclops aspericornis for the control of dengue vector,Aedes aegypti

Objective

To evaluate the toxicity, predatory efficiency of Delonix elata (D. elata) and Mesocyclops aspericornis (M. aspericornis) against dengue vector, Aedes aegypti (Ae. aegypti).

Methods

A mosquitocidal bioassay was conducted at different concentration of plant extract followed by WHO standard method. The probit analysis of each tested concentration and control were observed by using software SPSS 11 version package. The each tested concentration variable was assessed by DMRT method. The predatory efficiency of copepod was followed by Deo et al., 1988. The predator, M. aspericornis was observed for mortality, abnormalities, survival and swimming activity after 24 h treatment of plant and also predation on the mosquito larvae were observed.

Results

D. elata were tested for biological activity against the larvae, and pupae of Ae. aegypti. Significant mortality effects were observed in each life stage. The percentage of mortality was 100% in first and second instars whereas 96%, 92% in third and fourth instars. Fitted probit-mortality curves for larvae indicated the median and 90% lethal concentrations of D. elata for instars 1-4 to be 4.91 (8.13), 5.16 (8.44), 5.95 (7.76) and 6.87 (11.23), respectively. The results indicate that leaf extract exhibits significant biological activity against life stages. The present study revealed that D. elata is potentially important in the control of Ae. aegypti. Similar studies were conducted for predatory efficiency of Copepod, M. aspericornis against mosquito vector Ae. Aegypti. This study reported that the predatory copepod fed on 39% and 25% in I and III instar larvae of mosquito and in combined treatment of D. elata and copepod maximum control of mosquito larval states and at 83%, 80%, 75% and 53% in I, II, III and IV instars, respectively.

Conclusions

The combined action of plant extract and predatory copepod to effectively control mosquito population and reduce the dengue transmitting diseases.     

Modular columns to study depth-dependence behavior of mosquito larvae and toxicity of Bacillus thuringiensis subsp. israelensis

Modular transparent column system was designed to study depth-dependence behavior of mosquito larvae. The system was used in preliminary experiments to evaluate the effect of water depth on the larvicidal activity of Bacillus thuringiensis subsp. israelensis de Barjac against bottom feeder larvae of Aedes aegypti (Linn.) (Diptera: Culicidae), and suggestions for increasing the efficiency of the device are discussed.