Erik M. Blosser

The effects of interspecific courtship on the mating success of Aedes aegypti and Aedes albopictus (Diptera: Culicidae) males.

ABSTRACT Satyrization, a form of asymmetric reproductive interference, has recently been shown to play a role in competitive displacements of Aedes aegypti (L.) by Aedes albopictus (Skuse). Furthermore, female Ae. aegypti from populations in sympatry with Ae. albopictus have evolved reproductive character displacement and changes in mating behavior to reduce interspecific mating. In this article, we examine evolutionary responses of males to interspecific mating and show that satyrization has also evoked reproductive character displacement in males. We demonstrate that the presence of heterospecific females negatively influences conspecific mating success in male Ae. aegypti, most likely due to misdirected courting or mating efforts, and that males of this species from populations in sympatry with Ae. albopictus have evolved to be less influenced by the presence of heterospecific females than their allopatric counterparts. Conversely, we suggest that the presence of conspecifics may, in some circumstances, increase interspecific mating. This study demonstrates that co-occurrences of these two invasive species may lead to evolution and adaptation of reproductive behaviors to changing circumstances. Understanding the processes driving development of mate choice preferences or avoidance mechanisms may help predict future changes in the distribution and abundance of insect vectors or pests.

Host use and seasonality of Culex (Melanoconion) iolambdis (Diptera: Culicidae) from eastern Florida, USA.

Culex (Melanoconion) iolambdis (Dyar, 1918) is a mosquito (Diptera: Culicidae) species found throughout much of tropical America, including southern Florida. Relatively few reports are available regarding the ecology of Cx. iolambdis, despite its widespread distribution and putative involvement in transmission of Venezuelan equine encephalitis virus. To quantify habitat and host utilization, adults of Cx. iolambdis were sampled from resting shelters at a field site in Vero Beach, Florida, over a 12-month period. Culex iolambdis (1109 males, 3072 females) constituted more than half (56.76%) of all mosquitoes sampled (24 species) and was active year-round. Unfed females and gravid females of Cx. iolambdis were significantly more abundant in mangrove habitat, while males and blood-fed females were not. PCR-based bloodmeal analysis of 305 females revealed that Cx. iolambdis has very wide host breadth, feeding on birds (37.0% overall), reptiles (26.6%), amphibians (23.3%) and mammals (13.1%). Green heron (Butorides virescens), Southern leopard frog (Lithobates sphenocephala) and American alligator (Alligator mississippiensis) were the most commonly fed upon hosts. Bloodmeals from different host classes varied significantly with season, suggesting that Cx. iolambdis may play a role in the amplification and epidemic transmission of zoonotic arboviruses affecting human health.

Environmental Drivers of Seasonal Patterns of Host Utilization by Culiseta melanura (Diptera: Culicidae) in Florida.

Field studies were carried out in four Florida counties to investigate winter and spring ecology of host use by Culiseta melanura (Coquillet), the primary vector of eastern equine encephalomyelitis virus (EEEV) in North America. Bloodmeal analysis by PCR was used to identify 233 host bloodmeals, which mainly originated from birds (78.5%) and reptiles (17.2%), primarily Anolis spp. lizards. Across counties, the percentage of bloodmeals from reptiles (7-37% depending upon county) increased with increasing day length and temperature in the spring. Multiple logistic regression revealed that differences in reptile host use across collection sites were largely explained by differences in average day length and temperature on the day of collection, and is probably owing to environment-driven behavioral patterns of ectothermic animals. Although past studies have demonstrated reptile biting by epizootic vectors of EEEV, including Culex (Melanoconion) spp., this is the first study to demonstrate widespread and common feeding upon ectothermic hosts by Cs. melanura. This work suggests that reptiles, particularly anole lizards, play a role in the ecology of EEEV in Florida either as amplifying hosts or as noncompetent hosts which dilute vector feedings thereby suppressing transmission. Detailed laboratory studies investigating impacts of environmental variables (temperature and photoperiod) on EEEV competence of anoles are needed to assess whether these animals support virus amplification.

Carbon dioxide generated from carbonates and acids for sampling blood-feeding arthropods

Carbon dioxide (CO2) is utilized to attract mosquitoes and other blood-feeding arthropods to traps around the world. Commercial forms of CO2 (e.g., dry ice and compressed gas) are often unavailable or extremely expensive in developing nations, where vector surveillance is essential to make life-saving decisions. We developed and tested inexpensive and reproducible methods of CO2 production from the combination of acids and carbonates, ranging from very basic (crushed seashells and vinegar) to relatively elaborate (a device that controls the timing of the acid-carbonate reaction and extends the reaction over several hours). When utilized with mosquito traps in Florida, USA and black fly traps in Region des Cascades, Burkina Faso, these carbonate-acid CO2 sources attracted significantly greater numbers of both vector groups, than did unbaited traps. CO2 was generated for more than four hours at levels sufficient to attract vectors over the entire period. The utility of this simple methodology in developing nations should be further evaluated.

Unbiased Strain-Typing of Arbovirus Directly from Mosquitoes Using Nanopore Sequencing: A Field-forward Biosurveillance Protocol

The future of infectious disease surveillance and outbreak response is trending towards smaller hand-held solutions for point-of-need pathogen detection. Here, samples of Culex cedecei mosquitoes collected in Southern Florida, USA were tested for Venezuelan Equine Encephalitis Virus (VEEV), a previously-weaponized arthropod-borne RNA-virus capable of causing acute and fatal encephalitis in animal and human hosts. A single 20-mosquito pool tested positive for VEEV by quantitative reverse transcription polymerase chain reaction (RT-qPCR) on the Biomeme two3. The virus-positive sample was subjected to unbiased metatranscriptome sequencing on the Oxford Nanopore MinION and shown to contain Everglades Virus (EVEV), an alphavirus in the VEEV serocomplex. Our results demonstrate, for the first time, the use of unbiased sequence-based detection and subtyping of a high-consequence biothreat pathogen directly from an environmental sample using field-forward protocols. The development and validation of methods designed for field-based diagnostic metagenomics and pathogen discovery, such as those suitable for use in mobile “pocket laboratories”, will address a growing demand for public health teams to carry out their mission where it is most urgent: at the point-of-need.

Mammal decline, linked to invasive Burmese python, shifts host use of vector mosquito towards reservoir hosts of a zoonotic disease

Invasive apex predators have profound impacts on natural communities, yet the consequences of these impacts on the transmission of zoonotic pathogens are unexplored. Collapse of large- and medium-sized mammal populations in the Florida Everglades has been linked to the invasive Burmese python, Python bivittatus Kuhl. We used historic and current data to investigate potential impacts of these community effects on contact between the reservoir hosts (certain rodents) and vectors of Everglades virus, a zoonotic mosquito-borne pathogen that circulates in southern Florida. The percentage of blood meals taken from the primary reservoir host, the hispid cotton rat, Sigmodon hispidus Say and Ord, increased dramatically (422.2%) from 1979 (14.7%) to 2016 (76.8%), while blood meals from deer, raccoons and opossums decreased by 98.2%, reflecting precipitous declines in relative abundance of these larger mammals, attributed to python predation. Overall species diversity of hosts detected in Culex cedecei blood meals from the Everglades declined by 40.2% over the same period (H(1979) = 1.68, H(2016) = 1.01). Predictions based upon the dilution effect theory suggest that increased relative feedings upon reservoir hosts translate into increased abundance of infectious vectors, and a corresponding upsurge of Everglades virus occurrence and risk of human exposure, although this was not tested in the current study. This work constitutes the first indication that an invasive predator can increase contact between vectors and reservoirs of a human pathogen and highlights unrecognized indirect impacts of invasive predators.

Identification of Uranotaenia sapphirina as a specialist of annelids broadens known mosquito host use patterns

Feeding upon vertebrate blood by mosquitoes permits transmission of diverse pathogens, including viruses, protozoa, and nematodes. Despite over a century of intensive study, no mosquito species is known to specialize on non-vertebrate hosts. Using molecular analyses and field observations, we provide the first evidence, to our knowledge, that a mosquito, Uranotaenia sapphirina, specializes on annelid hosts (earthworms and leeches) while its sympatric congener, Uranotaenia lowii, feeds only on anurans (frogs and toads). Our results demonstrate that Ur. sapphirina feeds on annelid hosts (100% of identified blood meals; n = 72; collected throughout Florida), findings that are supported by field observations of these mosquitoes feeding on Sparganophilus worms and freshwater leeches. These findings indicate that adult mosquitoes utilize a much broader range of host taxa than previously recognized, with implications for epidemiology and the evolution of host use patterns in mosquitoes.Lawrence Reeves et al. report evidence that adult females of the mosquito species Uranotaenia sapphirina feed primarily on annelid hosts. This is the first known example of a mosquito species that specializes on invertebrate blood and suggests that mosquito host use patterns are more diverse than previously recognized.

Aedeomyia squamipennis (Diptera: Culicidae) in Florida, USA, a New State and Country Record

Abstract Aedeomyia squamipennis (Lynch Arribalzaga) is a tropical mosquito (Diptera: Culicidae) found throughout most of the American Tropics, from eastern Mexico through Argentina, including several Caribbean islands. Larvae are typically associated with bodies of water with dense growths of aquatic vegetation, particularly Pistia stratiotes L., water lettuce. Adult females feed predominantly on the blood of birds and seek hosts in forest canopies. Aedeomyia squamipennis is considered an important vector of Gamboa virus and avian malaria, and is also suspected of transmitting Venezuelan equine encephalitis virus. Collections from Florida City, FL, near the southern tip of the Florida peninsula, yielded larvae, adult males, and females of Ad. squamipennis, constituting a new genus and species record for Florida and the United States. The widespread availability of larval habitat and suitable hosts in Florida will likely lead to expansion of Ad. squamipennis in Florida, and perhaps into neighboring states. In South America, Ad. squamipennis is found as far south as Buenos Aires, Argentina, at ~35° S latitude, which is equivalent in N latitude to coastal North Carolina. The northern limit of expansion of Ad. squamipennis in North America will likely be limited by winter temperatures and availability of larval habitat.

Culex (Melanoconion) panocossa from peninsular Florida, USA

Culex (Melanoconion) panocossa is a suspected vector of Venezuelan equine encephalitis virus in Central America. Prior to this report, Cx. panocossa was known from Central America (Belize, Costa Rica, El Salvador, Guatemala, southern Mexico, Panama), northern South America (Colombia, Venezuela) and the Greater Antilles (Cuba and Jamaica). Larvae (n=5) and adults (n=4286) of Cx. panocossa were collected at two locations near Homestead, FL, which indicates substantial established populations of this probable vector species in the continental US. Since larvae of Cx. panocossa are associated with Pistia spp. (water lettuce), the distribution of this mosquito is likely to expand in Florida, where water lettuce is a major invasive plant in freshwater ecosystems. The putative establishment of Cx. panocossa in Florida is of significant concern from a public health perspective, as its proliferation in developed areas could link historically sylvatic transmission foci of Everglades virus with populated centers such as the greater Miami Metropolitan area.

Oviposition Strategies of Florida Culex (Melanoconion) Mosquitoes

Abstract In the genus Culex, oviposition is often generalized as laying floating egg rafts directly on the surface of the water, as reported for arbovirus vectors Culex pipiens L. and Culex tarsalis Coquillett, both members of subgenus Culex. The current work reports on investigations of exceptions to this strategy by species of Melanoconion, a diverse and mostly tropical subgenus of Culex, with many arbovirus vectors. Wild gravid females from Vero Beach, FL, were introduced into outdoor cages with open water, partially covered water, and emergent solid structures for oviposition. Three species, Culex cedecei Stone and Hair, Culex iolambdis Dyar, and Culex pilosus Dyar and Knab laid vast majorities (80–100%) of egg clusters attached to solid surfaces, although at different heights and angles, while Culex atratus Theobald laid floating egg clusters (100%) directly on the water. When offered mud, open water, and woody material, Cx. iolambdis laid 86% of clusters on nearly vertical surfaces of mangrove rootlets. Culex iolambdis eggs laid on solid surfaces hatched within 3–4 d or dried up without hatching. Results suggest that Melanoconion species utilize a variety of oviposition strategies, yet most species studied oviposit out of water. For some species, such as Cx. pilosus, this is a strategy for diapause and desiccation resistance, while for others, such as Cx. iolambdis, ovipositing above water may protect eggs from predators or being flushed from larval habitat. A summary of oviposition strategies in genus Culex shows that above-water oviposition is widespread and that the Culex egg raft paradigm needs reassessment.