Multiplex Surveillance Kit Using Sweetened Solid Support to Detect Arboviruses Isothermally in Mosquito Saliva from the Field.

Recently reported "displaceable probe" loop amplification (DP-LAMP) architecture has shown to amplify viral RNA from SARS-CoV-2 with little sample processing. The architecture allows signals indicating the presence of target nucleic acids to be spatially separated, and independent in sequence, from the complicated concatemer that LAMP processes create as part of their amplification process. This makes DP-LAMP an attractive molecular strategy to integrate with trap and sampling innovations to detect RNA from arboviruses carried by mosquitoes in the field. These innovations include (a) development of organically produced carbon dioxide with ethylene carbonate as a bait deployable in mosquito trap, avoiding the need for dry ice, propane tanks, or inorganic carbonates and (b) a process that induces mosquitoes to lay virus-infected saliva on a quaternary ammonium-functionalized paper (Q-paper) matrix, where (c) the matrix (i) inactivates the deposited viruses, (ii) releases their RNA, and (iii) captures viral RNA in a form that keeps it stable for days at ambient temperatures. We report this integration here, with a surprisingly simple workflow. DP-LAMP with a reverse transcriptase was found to amplify arboviral RNA directly from Q-paper, without requiring a separate elution step. This capture-amplification-detection architecture can be multiplexed, with the entire system integrated into a device that can support a campaign of surveillance, in the wild outdoors, that reports the prevalence of arboviruses from field-captured mosquitoes.

Patterns of Abundance, Host Use, and Everglades Virus Infection in Culex (Melanoconion) cedecei Mosquitoes, Florida, USA.

Everglades virus (EVEV), subtype II within the Venezuelan equine encephalitis (VEE) virus complex, is a mosquitoborne zoonotic pathogen endemic to south Florida, USA. EVEV infection in humans is considered rare, probably because of the sylvatic nature of the vector, the Culex (Melanoconion) cedecei mosquito. The introduction of Cx. panocossa, a tropical vector mosquito of VEE virus subtypes that inhabits urban areas, may increase human EVEV exposure. Field studies investigating spatial and temporal patterns of abundance, host use, and EVEV infection of Cx. cedecei mosquitoes in Everglades National Park found that vector abundance was dynamic across season and region. Rodents, particularly Sigmodon hispidus rats, were primary vertebrate hosts, constituting 77%-100% of Cx. cedecei blood meals. Humans were fed upon at several locations. We detected EVEV infection in Cx. cedecei mosquitoes in lower and upper regions of Everglades National Park only during the wet season, despite an abundance of Cx. cedecei mosquitoes at other sampling times.

Multiplexed kit based on Luminex technology and achievements in synthetic biology discriminates Zika, chikungunya, and dengue viruses in mosquitoes.

BACKGROUND: The global expansion of dengue (DENV), chikungunya (CHIKV), and Zika viruses (ZIKV) is having a serious impact on public health. Because these arboviruses are transmitted by the same mosquito species and co-circulate in the same area, a sensitive diagnostic assay that detects them together, with discrimination, is needed. METHODS: We present here a diagnostics panel based on reverse transcription-PCR amplification of viral RNA and an xMap Luminex architecture involving direct hybridization of PCRamplicons and virus-specific probes. Two DNA innovations ("artificially expanded genetic information systems", AEGIS, and "self-avoiding molecular recognition systems", SAMRS) increase the hybridization sensitivity on Luminex microspheres and PCR specificity of the multiplex assay compared to the standard approach (standard nucleotides). RESULTS: The diagnostics panel detects, if they are present, these viruses with a resolution of 20 genome equivalents (DENV1), or 10 (DENV3-4, CHIKV) and 80 (DENV2, ZIKV) genome equivalents per assay. It identifies ZIKV, CHIKV and DENV RNAs in a single infected mosquito, in mosquito pools comprised of 5 to 50 individuals, and mosquito saliva (ZIKV, CHIKV, and DENV2). Infected mosquitoes and saliva were also collected on a cationic surface (Q-paper), which binds mosquito and viral nucleic acids electrostatically. All samples from infected mosquitoes displayed only target-specific signals; signals from non-infected samples were at background levels. CONCLUSIONS: Our results provide an efficient and multiplex tool that may be used for surveillance of emerging mosquito-borne pathogens which aids targeted mosquito control in areas at high risk for transmission.

Linking nutrient stoichiometry to Zika virus transmission in a mosquito.

Food quality and quantity serve as the basis for cycling of key chemical elements in trophic interactions; yet the role of nutrient stoichiometry in shaping host-pathogen interactions is under appreciated. Most of the emergent mosquito-borne viruses affecting human health are transmitted by mosquitoes that inhabit container systems during their immature stages, where allochthonous input of detritus serves as the basal nutrients. Quantity and type of detritus (animal and plant) were manipulated in microcosms containing newly hatched Aedes aegypti mosquito larvae. Adult mosquitoes derived from these microcosms were allowed to ingest Zika virus-infected blood and then tested for disseminated infection, transmission, and total nutrients (percent carbon, percent nitrogen, ratio of carbon to nitrogen). Treatments lacking high-quality animal (insect) detritus significantly delayed development. Survivorship to adulthood was closely associated with the amount of insect detritus present. Insect detritus was positively correlated with percent nitrogen, which affected Zika virus infection. Disseminated infection and transmission decreased with increasing insect detritus and percent nitrogen. We provide the first definitive evidence linking nutrient stoichiometry to arbovirus infection and transmission in a mosquito using a model system of invasive Ae. aegypti and emergent Zika virus.

Transcriptional Profile of Aedes aegypti Leucine-Rich Repeat Proteins in Response to Zika and Chikungunya Viruses.

Aedes aegypti (L.) is the primary vector of chikungunya, dengue, yellow fever, and Zika viruses. The leucine-rich repeats (LRR)-containing domain is evolutionarily conserved in many proteins associated with innate immunity in invertebrates and vertebrates, as well as plants. We focused on the AaeLRIM1 and AaeAPL1 gene expressions in response to Zika virus (ZIKV) and chikungunya virus (CHIKV) infection using a time course study, as well as the developmental expressions in the eggs, larvae, pupae, and adults. RNA-seq analysis data provided 60 leucine-rich repeat related transcriptions in Ae. aegypti in response to Zika virus (Accession number: GSE118858, accessed on: August 22, 2018, GEO DataSets). RNA-seq analysis data showed that AaeLRIM1 (AAEL012086-RA) and AaeAPL1 (AAEL009520-RA) were significantly upregulated 2.5 and 3-fold during infection by ZIKV 7-days post infection (dpi) of an Ae. aegypti Key West strain compared to an Orlando strain. The qPCR data showed that LRR-containing proteins related genes, AaeLRIM1 and AaeAPL1, and five paralogues were expressed 100-fold lower than other nuclear genes, such as defensin, during all developmental stages examined. Together, these data provide insights into the transcription profiles of LRR proteins of Ae. aegypti during its development and in response to infection with emergent arboviruses.

Transcriptomic Analysis of Aedes aegypti Innate Immune System in Response to Ingestion of Chikungunya Virus.

Aedes aegypti (L.) is the primary vector of emergent mosquito-borne viruses, including chikungunya, dengue, yellow fever, and Zika viruses. To understand how these viruses interact with their mosquito vectors, an analysis of the innate immune system response was conducted. The innate immune system is a conserved evolutionary defense strategy and is the dominant immune system response found in invertebrates and vertebrates, as well as plants. RNA-sequencing analysis was performed to compare target transcriptomes of two Florida Ae. aegypti strains in response to chikungunya virus infection. We analyzed a strain collected from a field population in Key West, Florida, and a laboratory strain originating from Orlando. A total of 1835 transcripts were significantly expressed at different levels between the two Florida strains of Ae. aegypti. Gene Ontology analysis placed these genes into 12 categories of biological processes, including 856 transcripts (up/down regulated) with more than 1.8-fold (p-adj (p-adjust value) ≤ 0.01). Transcriptomic analysis and q-PCR data indicated that the members of the AaeCECH genes are important for chikungunya infection response in Ae. aegypti. These immune-related enzymes that the chikungunya virus infection induces may inform molecular-based strategies for interruption of arbovirus transmission by mosquitoes.

Mosquito responses to trait- and density-mediated interactions of predation.

Mosquito and predatory larvae often share the same habitat. Predators may influence mosquito prey populations through both lethal effect and non-lethal pathways. A series of experimental manipulations were used to distinguish between lethal (density-mediated interaction) and non-lethal (trait-mediated interaction) effects in a model system comprised of invasive prey mosquito, Aedes aegypti, and a predatory mosquito Toxorhynchites rutilus. Treatments with predators present or manipulations mimicking daily mortality (density reduction) reduced developmental time and recruitment to the adult stage. Daily records of adult survival of A. aegypti showed that exposure to predators during the juvenile stage shortened the lifespan of adults. This was also observed in treatments, where A. aegypti were replaced at the rate of consumption by T. rutilus. In contrast, numerical reductions in A. aegypti that mimicked daily rate of predation led to adults with the longest lifespan. These observations suggest strong effects of density and trait-mediated interactions in the influence of predators on mosquito biology relevant to their ability to transmit pathogens. These results have potentially important implications for disease control strategies. The primary approach to reduce risk of mosquito-borne diseases is through population reduction of the vectors. We show an unanticipated benefit of biological control by predation for the control of juvenile stages of mosquitoes. Specifically, mosquitoes that are exposed to predators but survive to adulthood will have compromised life expectancy, a key parameter in determining risk of disease transmission.

Point of sampling detection of Zika virus within a multiplexed kit capable of detecting dengue and chikungunya.

BACKGROUND: Zika, dengue, and chikungunya are three mosquito-borne viruses having overlapping transmission vectors. They cause diseases having similar symptoms in human patients, but requiring different immediate management steps. Therefore, rapid (< one hour) discrimination of these three viruses in patient samples and trapped mosquitoes is needed. The need for speed precludes any assay that requires complex up-front sample preparation, such as extraction of nucleic acids from the sample. Also precluded in robust point-of-sampling assays is downstream release of the amplicon mixture, as this risks contamination of future samples that will give false positives. METHODS: Procedures are reported that directly test urine and plasma (for patient diagnostics) or crushed mosquito carcasses (for environmental surveillance). Carcasses are captured on paper samples carrying quaternary ammonium groups (Q-paper), which may be directly introduced into the assay. To avoid the time and instrumentation requirements of PCR, the procedure uses loop-mediated isothermal amplification (LAMP). Downstream detection is done in sealed tubes, with dTTP-dUTP mixtures in the LAMP with a thermolabile uracil DNA glycosylase (UDG); this offers a second mechanism to prevent forward contamination. Reverse transcription LAMP (RT-LAMP) reagents are distributed dry without requiring a continuous chain of refrigeration. RESULTS: The tests detect viral RNA in unprocessed urine and other biological samples, distinguishing Zika, chikungunya, and dengue in urine and in mosquitoes infected with live Zika and chikungunya viruses. The limits of detection (LODs) are ~0.71 pfu equivalent viral RNAs for Zika, ~1.22 pfu equivalent viral RNAs for dengue, and ~38 copies of chikungunya viral RNA. A handheld, battery-powered device with an orange filter was constructed to visualize the output. Preliminary data showed that this architecture, working with pre-prepared tubes holding lyophilized reagent/enzyme mixtures and shipped without a chain of refrigeration, also worked with human plasma samples to detect chikungunya and dengue in Pune, India. CONCLUSIONS: A kit, complete with a visualization device, is now available for point-of-sampling detection of Zika, chikungunya, and dengue. The assay output is read in ca. 30 min by visualizing (human eye) three-color coded fluorescence signals. Assay in dried format allows it to be run in low-resource environments.

Dengue serotype-specific immune response in Aedes aegypti and Aedes albopictus.

BACKGROUND: Dengue viruses (DENV) are considered one of the most important emerging pathogens and dengue disease is a global health threat. The geographic expansion of dengue viruses has led to co-circulation of all four dengue serotypes making it imperative that new DENV control strategies be devised. OBJECTIVES: Here we characterize dengue serotype-specific innate immune responses in Aedes aegypti and Aedes albopictus using DENV from Puerto Rico (PR). METHODS: Ae. aegypti and Ae. albopictus were infected with dengue serotype 1 and 2 isolated from Puerto Rico. DENV infected mosquito samples were collected and temporal change in expression of selected innate immune response pathway genes analyzed by quantitative real time PCR. FINDINGS: The Toll pathway is involved in anti-dengue response in Ae. aegypti, and Ae. albopictus. Infections with PR DENV- 1 elicited a stronger response from genes of the Toll immune pathway than PR DENV-2 in Ae. aegypti but in infected Ae. albopictus expression of Toll pathway genes tended to be similar between the serotypes. Two genes (a ribosomal S5 protein gene and a nimrod-like gene) from Ae. albopictus were expressed in response to DENV. MAIN CONCLUSIONS: These studies revealed a role for antiviral genes in DENV serotype-specific interactions with DENV vectors, demonstrated that infections with DENV-2 can modulate the Toll immune response pathway in Ae. aegypti and elucidated candidate molecules that might be used to interfere with serotype specific vector-virus interactions.

Larval Temperature-Food Effects on Adult Mosquito Infection and Vertical Transmission of Dengue-1 Virus.

Temperature-food interactions in the larval environment can affect life history and population growth of container mosquitoes Aedes aegypti (L.) and Aedes albopictus Skuse, the primary vectors of chikungunya and dengue viruses. We used Ae. aegypti, Ae. albopictus, and dengue-1 virus (DENV-1) from Florida to investigate whether larval rearing temperature can alter the effects of larval food levels on Ae. aegypti and Ae. albopictus life history and DENV-1 infection and vertical transmission. Although we found no effect of larval treatments on survivorship to adulthood, DENV-1 titer, or DENV-1 vertical transmission, rates of vertical transmission up to 16-24% were observed in Ae. albopictus and Ae. aegypti, which may contribute to maintenance of this virus in nature. Larval treatments had no effect on number of progeny and DENV-1 infection in Ae. aegypti, but the interaction between temperature and food affected number of progeny and DENV-1 infection of the female Ae. albopictus parent. The cooler temperature (24°C) yielded the most progeny and this effect was accentuated by high food relative to the other conditions. Low and high food led to the highest (∼90%) and lowest (∼65%) parental infection at the cooler temperature, respectively, whereas intermediate infection rates (∼75-80%) were observed for all food conditions at the elevated temperature. These results suggest that temperature and food availability have minimal influence on rate of vertical transmission and a stronger influence on adults of Ae. albopictus than of Ae. aegypti, which could have consequences for dengue virus epidemiology.

Testing of Visual and Chemical Attractants in Correlation with the Development and Field Evaluation of an Autodissemination Station for the Suppression of Aedes aegypti and Aedes albopictus in Florida.

Three exotic mosquito-borne pathogens-dengue, chikungunya, and Zika viruses-transmitted by Aedes albopictus and Ae. aegypti have undergone dramatic global expansion in recent years. The control of vector populations and minimizing bites from these vectors are the primary methods of reducing risk of transmission of these viruses to humans. However, Ae. albopictus and Ae. aegypti are notoriously challenging to control through conventional chemical means, due primarily to difficulties in applying pesticides to their cryptic larval habitats. A novel strategy for suppressing populations of these species is the autodissemination of insect growth regulators (IGRs), in which adult female mosquitoes are attracted to a treatment station where they are tainted with small amounts of potent IGR. When the adult females subsequently visit oviposition sites, they inadvertently disseminate the IGR to larval development sites, suppressing their own population. Implementing this technology to control natural vector populations presents substantial logistical challenges. The current manuscript describes laboratory bioassays and field evaluations to design a novel autodissemination station (ADS) and test the methodology at field locations in Florida where Ae. aegypti and Ae. albopictus are abundant and pose a risk for transmission of emerging pathogens. The prototype ADS is intended to attract host-seeking, resting site-seeking, and oviposition site-seeking females through a combination of visual and olfactory cues. The efficacy of this strategy was assessed through the use of sentinel ovicups at field locations in Indian River County and Martin County, FL. Greatest efficacy (45.3 ± 7.7% mortality in treatment sentinel ovicups) was achieved at a field site with few competing natural ovisites, while much lower efficacy was observed in locations with numerous competing ovisites (0.0 to 29.0 ± 8.2% mortality). The efficacy of the ADS is likely to be strongly affected by the abundance of competing ovisites, the population dynamics, and climatic conditions.

Interspecific Larval Competition Differentially Impacts Adult Survival in Dengue Vectors.

Mosquitoes often experience intraspecific and interspecific competition among larvae attributable to high densities and nutrient limitation, especially container mosquitoes including Aedes aegypti (L.) and Aedes albopictus (Skuse). Density-dependent effects on larvae impact adult production and adult traits that influence transmission of arboviruses. To improve our understanding of the mechanisms by which density-dependence influences transmission and identify species-specific traits, we tested the hypotheses: 1) Competitive asymmetry in favor of Ae. albopictus over Ae. aegypti translates to altered adult female survival, and 2) Ae. aegypti adult females are more resistant to life-shortening effects of low-humidity conditions than Ae. albopictus. We gauged the relative impact of inter- and intraspecific larval competition on adult survival in high- and low-humidity regimes (77 and 44% relative humidity, respectively). For Ae. albopictus, intraspecific but not interspecific competition usually reduced adult survival under both humidity regimes. For Ae. aegypti, both intraspecific and interspecific competition reduced adult survival. Ae. albopictus adult survival was minimally influenced by interspecific competition with Ae. aegypti, consistent with observations that Ae. albopictus is the superior competitor. A species comparison indicated that Ae. aegypti exhibited a survival advantage relative to Ae. albopictus under both low- and high-humidity conditions. However, similar survival of these Aedes species was observed in some cases depending on conditions experienced in both the aquatic and terrestrial environments. These results demonstrate plasticity in survival rates of dengue and chikungunya vectors and the significance of considering the influence of biological interactions during the immature stages and abiotic conditions during the adult stage.

Trait- and density-mediated effects of predation on fecundity and fertility of Aedes aegypti (Diptera: Culicidae) mosquitoes.

Prey populations that encounter predators experience density-mediated effects through lethality and associated numerical changes in the population. Prey also exhibit trait-mediated effects through nonlethal alterations in phenotypic traits associated with exposure to predators. Immature stages of mosquitoes commonly co-occur in habitats along with predators, a natural source of mortality and potential biocontrol agent. Toxorhynchites rutilus Coquillett 1896 is a natural source of mortality with potential as a biological control agent. Previous studies have shown that predation and the mere presence of Tx. rutilus (predator cues) can alter the life-history traits of Aedes aegypti (L. 1762). In addition to observed reductions in recruitment of adults (lethality), exposure to Tx. rutilus without consumption resulted in adult Ae. aegypti females with altered growth and reduced lifespan. To determine the influence of predation on the reproductive biology of Ae. aegypti, we tested the hypothesis that predation, or exposure to predator cues, will compromise the reproduction of adult survivors through reductions in fecundity (egg batch size) and fertility (hatch rate). We observed that for both female and male Ae. aegypti, survival to adulthood was the lowest and development time the shortest in treatments containing prey removal effects, attributable to predation and treatment manipulations of density reduction. There were effects of Ae. aegypti weight, but not predation treatments, on fecundity and fertility. Results suggest that predator-mediated effects on Ae. aegypti derive from lethal effects due to consumption and alterations in other phenotypic traits of survivors, including development, weight, lifespan of adults, and population growth, but not reproductive parameters measured here.

Does prior exposure to larvicides influence dengue virus susceptibility in Aedes aegypti (Diptera: Culicidae)?

Control of mosquito vector populations is primarily intended to reduce the transmission of pathogens they transmit. Use of chemical controls, such as larvicides, can have unforeseen consequences on adult traits if not applied properly. The consequences of under application of larvicides are little studied, specifically the impacts on pathogen infection and transmission by the vectors that survive exposure to larvicides. We compared vector susceptibility of Aedes aegypti (L.) for dengue virus, serotype 1 (DENV-1) previously exposed as larvae to an LC50 of different classes of insecticides as formulated larvicides. Larval exposure to insect growth regulators (methoprene and pyriproxyfen) significantly increased susceptibility to infection of DENV-1 in Ae. aegypti adults but did not alter disseminated infection or transmission. Larval exposure to temephos, spinosad, and Bti did not increase infection, disseminated infection, or transmission of DENV-1. Our findings describe a previously under observed phenomenon, the latent effects of select larvicides on mosquito vector susceptibility for arboviruses. These data suggest that there are unintended consequences of sublethal exposure to select larvicides that can influence susceptibility of Ae. aegypti to DENV infection, and indicates the need for further investigation of sublethal effects of insecticides on other aspects of mosquito biology, especially those parameters relevant to a mosquitoes ability to transmit arboviruses (life span, biting behavior, extrinsic incubation period).

Rapid detection of West Nile and Dengue viruses from mosquito saliva by loop-mediated isothermal amplification and displaced probes.

Arthropod-borne viruses are major causes of human and animal disease, especially in endemic low- and middle-income countries. Mosquito-borne pathogen surveillance is essential for risk assessment and vector control responses. Sentinel chicken serosurveillance (antibody testing) and mosquito pool screening (by RT-qPCR or virus isolation) are currently used to monitor arbovirus transmission, however substantial time lags of seroconversion and/or laborious mosquito identification and RNA extraction steps sacrifice their early warning value. As a consequence, timely vector control responses are compromised. Here, we report on development of a rapid arbovirus detection system whereby adding sucrose to reagents of loop-mediated isothermal amplification with displaced probes (DP-LAMP) elicits infectious mosquitoes to feed directly upon the reagent mix and expectorate viruses into the reagents during feeding. We demonstrate that RNA from pathogenic arboviruses (West Nile and Dengue viruses) transmitted in the infectious mosquito saliva was detectable rapidly (within 45 minutes) without RNA extraction. Sucrose stabilized viral RNA at field temperatures for at least 48 hours, important for transition of this system to practical use. After thermal treatment, the DP-LAMP could be reliably visualized by a simple optical image sensor to distinguish between positive and negative samples based on fluorescence intensity. Field application of this technology could fundamentally change conventional arbovirus surveillance methods by eliminating laborious RNA extraction steps, permitting arbovirus monitoring from additional sites, and substantially reducing time needed to detect circulating pathogens.

Evolution of increased survival in RNA viruses specialized on cancer-derived cells.

Viruses and other pathogens can diverge in their evolved host-use strategies because of exposure to different host types and conflicts between within-host reproduction and between-host survival. Most host-pathogen studies have emphasized the role of intrahost reproduction in the evolution of pathogen virulence, whereas the role of extra-host survival has received less attention. Here, we examine the evolution of free-living virion survival in RNA virus populations differing in their histories of host use. To do so, we used lineages of vesicular stomatitis virus (VSV) that were experimentally evolved in laboratory tissue culture for 100 generations on cancer-derived cells, noncancerous cells, or alternating passages of the two host types. We observed that free-living survival improved when VSV populations specialized on human epithelial carcinoma (HeLa) cells, whereas this trait was not associated with selection on noncancer cells or combinations of the cell types. We attributed this finding to shorter-lived HeLa monolayers and/or rapid cell-to-cell spread of viruses on HeLa cells in tissue culture, both of which could select for enhanced virus stability between host-cell replenishment. We also showed evidence that increases in virion survival were associated with decreases in virulence, which suggests a trade-off between survival and virulence for the VSV populations on one cell type. Our results shed new light on the causes and consequences of "sit and wait" infection strategies in RNA viruses.

Vertical transmission of Key West dengue-1 virus by Aedes aegypti and Aedes albopictus (Diptera: Culicidae) mosquitoes from Florida.

Following the 2009 and 2010 dengue-1 (DENV-1) outbreaks in Key West, FL, we used Florida Aedes aegypti (L.) mosquitoes and DENV-1 isolated from Key West in 2010 to test the hypothesis that if the 2009 and 2010 DENV-1 genome sequences are similar, then vertical transmission of DENV-1 from infected Ae. aegypti female mosquitoes to their eggs could have served as an interepidemic reservoir between outbreaks. We also investigated the ability of Florida Aedes albopictus (Skuse) mosquitoes to vertically transmit DENV-1. In addition, we determined the rates of infection and dissemination of these Florida mosquito species for DENV-1 and the effect of DENV-1 infection on oviposition success and number of mosquito eggs laid by females. Vertical transmission of DENV-1 was documented, with rates of 11.11% (2 out of 18) for Ae. albopictus and 8.33% (3 out of 36) for Ae. aegypti. Approximately 93% (111 out of 119) of Ae. aegypti that fed on DENV-1 in blood became infected, and 80% (89 out of 111) of infections were disseminated. Similarly, 93% of Ae. albopictus became infected (53 out of 57), and 85% (45 out of 53) of infections were disseminated. No significant differences were detected in numbers of eggs laid by either species after imbibing DENV-1 in blood, suggesting little cost of infection on number of eggs laid. Our results demonstrate that Florida Ae. aegypti and Ae. albopictus mosquitoes are competent vectors for DENV-1, whose maintenance between the 2009 and 2010 Key West outbreaks may have been facilitated by vertical transmission.

Pesticide-induced release from competition among competing Aedes aegypti and Aedes albopictus (Diptera: Culicidae).

Competitive interactions between mosquitoes Aedes aegypti (L.) and Aedes albopictus (Skuse) may depend on environmental conditions. Pesticides may alleviate density-dependent competition for limited food, and a differential species response to sublethal concentrations may modify interspecific competition. We tested the hypothesis that exposure to malathion alters interspecific resource competition between these two species. In the absence of malathion, Ae. aegypti survivorship and per capita rate of population change were negatively affected by increasing densities of Ae. albopictus. However, the asymmetrical negative effect ofAe. albopictus on Ae. aegypti was eliminated in the presence of malathion. In addition, the presence of malathion resulted in shorter development time compared with the controls. The relative importance of pesticide-mediated coexistence in nature has not been evaluated, so its role in mediating coexistence is unclear; however, these findings underscore the potential of environmental concentrations of malathion, and perhaps other pesticides to facilitate coexistence between species.

West Nile virus infection rates and avian serology in east-central Illinois.

Understanding the geographic role of different species of mosquito vectors and vertebrate hosts in West Nile virus (WNV) transmission cycles can facilitate the development and implementation of targeted surveillance and control measures. This study examined the relationship between WNV-antibody rates in birds and mosquito infection rates and bloodfeeding patterns in east-central Illinois. The earliest detection of WNV-RNA by reverse transcription-polymerase chain reaction TaqMan was from Culex restuans; however, amplification typically coincided with an increase in abundance of Cx. pipiens. Trap type influenced annual estimates of infection rates in Culex species, as well as estimation of blood meal source. Bird species with the highest WNV-antibody rates (i.e., Mourning Doves [Zenaida macroura], Northern Cardinals [Cardinalis cardinalis], American Robins [Turdus migratorius], and House Sparrows [Passer domesticus]) were also the common species found in Culex blood meals. Although antibody rates were not directly proportional to estimated avian abundance, the apparent availability of mammal species did influence proportion of mammal to bird blood meals. Antibody prevalence in the American Robin was lower than expected based on the strong attraction of Culex to American Robins for blood meals. Age-related differences in serology were evident, antibody rates increased in older groups of robins and sparrows, whereas 1st-year hatch and older adults of Mourning Doves and Northern Cardinals had equally high rates of antibody-positive serum samples. The vector and host interactions observed in east-central Illinois (Champaign County), an urban area surrounded by agriculture, are compared to studies in the densely population areas of southern Cook County.

Transmission of Asian Zika Lineage by Aedes aegypti and Ae. albopictus Mosquitoes in Florida.

The Asian lineage of Zika virus (ZIKV), a mosquito-borne pathogen originally from Africa, caused an epidemic into Brazil in 2015 and subsequently spread throughout the Americas. Local transmission in the U.S. is a public health concern, especially for Florida where the mosquito vectors Aedes aegypti and Ae. albopictus are widespread, abundant, and there is a high potential for virus introduction due to imported cases. Here we evaluate relative susceptibility to infection and transmission of Zika virus among geographic populations of Ae. aegypti and Ae. albopictus in Florida. Both species have been implicated as ZIKV vectors elsewhere, but both virus and vector genotype are known to influence transmission capacities and, hence, the risk of outbreaks. We test the hypothesis that Ae. aegypti and Ae. albopictus show geographic differences in midgut and salivary gland barriers that limit ZIKV transmission, using local populations of the two vector species recently colonized from three regions of Florida to compare their susceptibility to ZIKV infection, disseminated infection, and transmission potential. Susceptibility to infection was higher in Ae. aegypti (range 76-92%) than Ae. albopictus (range 47-54%). Aedes aegypti exhibited 33-44% higher susceptibility to infection than Ae. albopictus, with Ae. aegypti from Okeechobee, FL having 17% higher susceptibility to infection than Ae. aegypti from Miami, FL. Similarly, disseminated infection was higher in Ae. aegypti (range 87-89%) than Ae. albopictus (range 31-39%), although did not vary by region. Enhanced infection and disseminated infection in Ae. aegypti were associated with higher viral loads in mosquito samples than in Ae. albopictus. Transmission rates did not vary by species or region (range 26-47%). The results support the hypothesis that Ae. aegypti, but not Ae. albopictus, exhibited regional differences in midgut infection barriers. Our observation of higher vector competence for Ae. aegypti than Ae. albopictus, together with this species greater propensity to feed on humans, lends support to the notion that Ae. aegypti is regarded as the primary vector for ZIKV and public health concern in continental U.S.