Utilization of a zoo for mosquito (Diptera: Culicidae) diversity analysis, arboviral surveillance, and blood feeding patterns.

Zoos provide a unique opportunity to study mosquito feeding ecology as they represent areas where exotic animals, free-roaming native animals, humans, and mosquito habitats overlap. Therefore, these locations are a concern for arbovirus transmission to both valuable zoo animals and human visitors. We sampled mosquitoes in and around The Nashville Zoo at Grassmere in Tennessee, USA, over 4 months in 2020 using 4 mosquito trap methods and 12 sampling locations. Mosquitoes were identified to species, Culex mosquitoes were analyzed for arboviruses, and all engorged mosquitoes were preserved for host usage analysis. We captured over 9,000 mosquitoes representing 27 different species, including a new species record for Davidson County, TN (Culex nigripalpus Theobald). Minimum infection rates for West Nile virus (WNV) (Flaviviridae: Flavivirus), St. Louis encephalitis virus (Flaviviridae: Flavivirus), and Flanders virus (Hapavirus: Rhabdoviridae) were 0.79, 0, and 4.17, respectively. The collection of 100 engorged mosquitoes was dominated by Culex pipiens pipiens Linnaeus (38%), Culex erraticus Dyar and Knab (23%), and Culex pipiens pipiens-Culex pipiens quinquefasciatus hybrids (10%). Host DNA from 84 engorged mosquitoes was successfully matched to a variety of host species (n = 23), with just 8 species belonging to the zoo. Wild birds were the most frequently fed upon host, in particular northern cardinals (Cardinalis cardinalis L. Passeriformes: Cardinalidae), which are competent WNV reservoirs. Taken together, our results demonstrate the utility of zoos as sentinels for emerging pathogens, for studying wildlife and human risk of zoonotic diseases, and for assessing vector diversity.

Aedes albopictus host odor preference does not drive observed variation in feeding patterns across field populations.

Laboratory and field-based studies of the invasive mosquito Aedes albopictus demonstrate its competency to transmit over twenty different pathogens linked to a broad range of vertebrate hosts. The vectorial capacity of Ae. albopictus to transmit these pathogens remains unclear, partly due to knowledge gaps regarding its feeding behavior. Blood meal analyses from field-captured specimens have shown vastly different feeding patterns, with a wide range of anthropophagy (human feeding) and host diversity. To address this knowledge gap, we asked whether differences in innate host preference may drive observed variation in Ae. albopictus feeding patterns in nature. Low generation colonies (F2-F4) were established with field-collected mosquitoes from three populations with high reported anthropophagy (Thailand, Cameroon, and Florida, USA) and three populations in the United States with low reported anthropophagy (New York, Maryland, and Virginia). The preference of these Ae. albopictus colonies for human versus non-human animal odor was assessed in a dual-port olfactometer along with control Ae. aegypti colonies already known to show divergent behavior in this assay. All Ae. albopictus colonies were less likely (p < 0.05) to choose the human-baited port than the anthropophilic Ae. aegypti control, instead behaving similarly to zoophilic Ae. aegypti. Our results suggest that variation in reported Ae. albopictus feeding patterns are not driven by differences in innate host preference, but may result from differences in host availability. This work is the first to compare Ae. albopictus and Ae. aegypti host preference directly and provides insight into differential vectorial capacity and human feeding risk.

The Effects of Host Availability and Fitness on Aedes albopictus Blood Feeding Patterns in New York.

Aedes albopictus is a competent vector of numerous pathogens, representing a range of transmission cycles involving unique hosts. Despite the important status of this vector, variation in its feeding patterns is poorly understood. We examined the feeding patterns of Ae. albopictus utilizing resting collections in Long Island, NY, and contextualized blood meal sources with host availability measured by household interviews and camera traps. We identified 90 blood meals, including 29 humans, 22 cats, 16 horses, 12 opossums, 5 dogs, 2 goats, and 1 each of rabbit, rat, squirrel, and raccoon. This is only the third study of Ae. albopictus blood feeding biology that quantitatively assessed domestic host availability and is the first to do so with wild animals. Host feeding indices showed that cats and dogs were fed upon disproportionately often compared with humans. Forage ratios suggested a tendency to feed on cats and opossums and to avoid raccoons, squirrels, and birds. This feeding pattern was different from another published study from Baltimore, where Ae. albopictus fed more often on rats than humans. To understand whether these differences were because of host availability or mosquito population variation, we compared the fitness of New York and Baltimore Ae. albopictus after feeding on rat and human blood. In addition, we examined fitness within the New York population after feeding on human, rat, cat, horse, and opossum blood. Together, our results do not indicate major mosquito fitness differences by blood hosts, suggesting that fitness benefits do not drive Northeastern Ae. albopictus feeding patterns.

Understanding and interpreting mosquito blood feeding studies: the case of Aedes albopictus.

Blood feeding is a fundamental mosquito behavior with consequences for pathogen transmission and control. Feeding behavior can be studied through two lenses - patterns and preference. Feeding patterns are assessed via blood meal analyses, reflecting mosquito-host associations influenced by environmental and biological parameters. Bias can profoundly impact results, and we provide recommendations for mitigating these effects. We also outline design choices for host preference research, which can take many forms, and highlight their respective (dis)advantages for preference measurement. Finally, Aedes albopictus serves as a case study for how to apply these lessons to interpret data and understand feeding biology. We illustrate how assumptions and incomplete evidence can lead to inconsistent interpretations by reviewing Ae. albopictus feeding studies alongside prevalent narratives about perceived behavior.

Sugar feeding patterns of New York Aedes albopictus mosquitoes are affected by saturation deficit, flowers, and host seeking.

BACKGROUND: Sugar feeding is an important behavior which may determine vector potential of female mosquitoes. Sugar meals can reduce blood feeding frequency, enhance survival, and decrease fecundity, as well as provide energetic reserves to fuel energy intensive behaviors such as mating and host seeking. Sugar feeding behavior can be harnessed for vector control (e.g. attractive toxic sugar baits). Few studies have addressed sugar feeding of Aedes albopictus, a vector of arboviruses of public health importance, including dengue and Zika viruses. To address this knowledge gap, we assessed sugar feeding patterns of Ae. albopictus for the first time in its invasive northeastern USA range. METHODOLOGY/PRINCIPAL FINDINGS: Using the cold anthrone fructose assay with robust sample sizes, we demonstrated that a large percentage of both male (49.6%) and female (41.8%) Ae. albopictus fed on plant or homopteran derived sugar sources within 24 hrs prior to capture. Our results suggest that sugar feeding behavior increases when environmental conditions are dry (high saturation deficit) and may vary by behavioral status (host seeking vs. resting). Furthermore, mosquitoes collected on properties with flowers (>3 blooms) had higher fructose concentrations compared to those collected from properties with few to no flowers (0-3). CONCLUSIONS/SIGNIFICANCE: Our results provide the first evidence of Ae. albopictus sugar feeding behavior in the Northeastern US and reveal relatively high rates of sugar feeding. These results suggest the potential success for regional deployment of toxic sugar baits. In addition, we demonstrate the impact of several environmental and mosquito parameters (saturation deficit, presence of flowers, host seeking status, and sex) on sugar feeding. Placing sugar feeding behavior in the context of these environmental and mosquito parameters provides further insight into spatiotemporal dynamics of feeding behavior for Ae. albopictus, and in turn, provides information for evidence-based control decisions.

Assessment of synthetic floral-based attractants and sugar baits to capture male and female Aedes aegypti (Diptera: Culicidae).

BACKGROUND: The viruses transmitted by Aedes aegypti, including dengue and Zika viruses, are rapidly expanding in geographic range and as a threat to public health. In response, control programs are increasingly turning to the use of sterile insect techniques resulting in a need to trap male Ae. aegypti to monitor the efficacy of the intervention. However, there is a lack of effective and cheap methods for trapping males. Thus, we attempted to exploit the physiological need to obtain energy from sugar feeding in order to passively capture male and female Ae. aegypti (nulliparous and gravid) in free-flight attraction assays. Candidate lures included previously identified floral-based (phenylacetaldehyde, linalool oxide, phenylethyl alcohol, and acetophenone) attractants and an attractive toxic sugar bait-based (ATSB) solution of guava and mango nectars. A free-flight attraction assay assessed the number of mosquitoes attracted to each candidate lure displayed individually. Then, a choice test was performed between the best-performing lure and a water control displayed in Gravid Aedes Traps (GAT). RESULTS: Results from the attraction assays indicated that the ATSB solution of guava and mango nectars was the most promising lure candidate for males; unlike the floral-based attractants tested, it performed significantly better than the water control. Nulliparous and gravid females demonstrated no preference among the lures and water controls indicating a lack of attraction to floral-based attractants and sugar baits in a larger setting. Although the guava-mango ATSB lure was moderately attractive to males when presented directly (i.e. no need to enter a trap or other confinement), it failed to attract significantly more male, nulliparous female, or gravid female Ae. aegypti than water controls when presented inside a Gravid Aedes Trap. CONCLUSIONS: Our findings suggest that the use of volatile floral-based attractants and sugar mixtures that have been identified in the literature is not an effective lure by which to kill Ae. aegypti at ATSB stations nor capture them in the GAT. Future trapping efforts would likely be more successful if focused on more promising methods for capturing male and female Ae. aegypti.

Evaluation of Alternative Killing Agents for Aedes aegypti (Diptera: Culicidae) in the Gravid Aedes Trap (GAT).

The Gravid Aedes Trap (GAT) uses visual and olfactory cues to attract gravid Aedes aegypti (L.) that are then captured when knocked down by a residual pyrethroid surface spray. However, the use of surface sprays can be compromised by poor availability of the spray and pesticide resistance in the target mosquito. We investigated several "alternative" insecticide and insecticide-free killing agents for use in the GAT. This included long-lasting insecticide-impregnated nets (LLINs), vapor-active synthetic pyrethroids (metofluthrin), canola oil, and two types of dry adhesive sticky card. During bench top assays LLINs, metofluthrin, and dry sticky cards had 24-h knockdown (KD) percentages >80% (91.2 ± 7.2%, 84.2 ± 6.8%, and 83.4 ± 6.1%, respectively), whereas the 24-h KD for canola oil was 70 ± 7.7%, which improved to 90.0 ± 3.7% over 48 h. Importantly, there were no significant differences in the number of Ae. aegypti collected per week or the number of traps positive for Ae. aegypti between the sticky card and canola oil treatments compared with the surface spray and LLIN treatments in semifield and field trials. These results demonstrate that the use of inexpensive and widely available insecticide-free agents such as those described in this study are effective alternatives to pyrethroids in regions with insecticide-resistant populations. The use of such environmentally friendly insecticide-free alternatives will also be attractive in areas where there is substantial resistance to insecticide use due to environmental and public health concerns.