Sustainable control of mosquito larvae in the field by the combined actions of the biological insecticide Bti and natural competitors

Integrated management of mosquitoes is becoming increasingly important, particularly in relation to avoiding recolonization of ponds after larvicide treatment. We conducted for the first time field experiments that involved exposing natural populations of the mosquito species Culex pipiens to: a) application of the biological insecticide Bacillus thuringiensis israelensis (Bti), b) the introduction of natural competitors (a crustacean community composed mainly of Daphnia spp.), or c) a combined treatment that involved both introduction of a crustacean community and the application of Bti. The treatment that involved only the introduction of crustaceans had no significant effect on mosquito larval populations, while treatment with Bti alone caused only a significant reduction in the abundance of mosquito larvae in the short‐term (within 3–10 days after treatment). In contrast, the combined treatment rapidly reduced the abundance of mosquito larvae, which remained low throughout the entire observation period of 28 days. Growth of the introduced crustacean communities was favored by the immediate reduction in the abundance of mosquito larvae following Bti administration, thus preventing recolonization of ponds by mosquito larvae at the late period (days 14–28 after treatment). Both competition and the temporal order of establishment of different species are hence important mechanisms for efficient and sustainable mosquito control.     

Temporal and spatial habitat preferences and biotic interactions between mosquito larvae and antagonistic crustaceans in the field

Investigations on natural antagonists of mosquito larvae found that micro‐crustaceans (e.g., Cladocera) control mosquito populations under experimental conditions. However, their relevance for mosquito control under field situations remains widely unclear because important information about habitat preferences and time of occurrence of crustaceans and mosquito larvae are still missing. In order to fill this knowledge gap, a field study was undertaken in different wetland areas of Saxony, Germany, in different habitats (i.e., grassland, forest, and reed‐covered wetlands). We found negative interactions between larvae of Ae. vexans and predatory Cyclopoida (Crustacean: Copepoda), which both were dominant during the first two weeks of hydroperiod, at ponds located at grassland habitats. Larvae of Cx. pipiens were spatially associated with competing Cladocera, but they colonized ponds more rapidly. Populations of Cladocera established from the third week of hydroperiod and prevented Cx. pipiens colonization thereafter. Ostracoda were highly abundant during the whole hydroperiod, but their presence was restricted to habitats of reed‐covered wetland at one geographical area. Mosquito larvae hardly occurred at those ponds. In general, we found that ponds at the reed‐covered wetlands provided better conditions for the initial development of crustaceans and hence, mosquito larval colonization was strongly inhibited. Grassland habitat, in contrast, favored early development of mosquito larvae. This study showed that micro‐crustaceans are relevant for mosquito management but their impact on mosquito larvae varies between species and depends on environmental conditions.     

Predation differentially structures immature mosquito populations in stormwater ponds

1. Understanding the factors underlying the abundance and distribution of species requires the consideration of a complex suite of interacting biotic and abiotic factors operating on multiple spatial and temporal scales. Larval mosquitoes inhabiting small human-constructed ponds represent a unique opportunity to investigate the relative importance of these structuring mechanisms while simultaneously generating applied knowledge on mosquito control. 2. A multi-year field survey of 32 stormwater ponds was conducted in central Illinois (Champaign County). From each pond, data were collected on pond structure type and hydroperiod, the presence/absence of cattails (Typha spp.), and measures of total nitrogen, phosphorus, and organic carbon, and chlorophyll a. The communities of crustacean zooplankton and aquatic insects were characterised, and these taxa were assigned into two main groups: predators and competitors of larval mosquitoes. Structural equation modelling was used to explore the direct and indirect effects of these biotic and abiotic factors on larval density for three species of culicine mosquitoes (Culex pipiens, Culex restuans, and Aedes vexans). 3. Hydroperiod had an indirect negative effect on C. pipiens density. However, this effect was mediated by predator density; more permanent ponds had more predators, which therefore reduced the density of C. pipiens larvae. Aedes vexans density was positively correlated with predator density. No predictor variables were found that explained variation in C. restuans density. 4. This study show that the relative importance of these biotic and abiotic factors varies among species of culicine mosquitoes inhabiting stormwater ponds.     

Bioinsecticide and leaf litter combination increases oviposition and reduces adult recruitment to create an effective ovitrap for Culex mosquitoes

Mosquito egg traps, aquatic habitats baited with oviposition attractant and insecticide, are important tools for surveillance and control efforts in integrated vector management programs. The bioinsecticide Bacillus thuringiensis subsp. israelensis (Bti) is increasingly used as an environmentally friendly alternative to chemical insecticides and the combination of Bti with a simple oviposition attractant like leaf litter to create an effective egg trap seems appealing. However, previous research suggests that Bti may itself alter oviposition, and that leaf litter may dramatically reduce Bti toxicity. Here we present results from field experiment designed to link the effects of litter and Bti on mosquito oviposition habitat selection and post‐colonization survival to production of adult mosquitoes. Tripling litter increased Culex spp. oviposition nearly nine‐fold, while Bti had no effect on oviposition. Neither factor altered egg survival, thus larval abundance reflected the effects of litter on oviposition. Both Bti and litter reduced larval survival by ～60%. We found no evidence that increased litter reduced Bti toxicity. Adult production was dependent upon both litter and Bti. In the absence of Bti, effects of litter on oviposition translated into three‐fold more adults. However, in the presence of Bti, initial increases in oviposition were erased by the combined negative effects of Bti and litter on post‐colonization survival. Thus, our study provides field evidence that combined litter and Bti application creates an effective ovitrap. This combined treatment had the highest oviposition and the lowest survival, and thus removed the greatest number of mosquitoes from the landscape.     

Controlling Culex pipiens: antagonists are more efficient than a neonicotinoid insecticide

Species vulnerability to pesticides depends on physiological sensitivity, the potential to recover, and the ecological context. We assessed the vulnerability of the mosquito Culex pipiens to a repeated treatment with thiacloprid in outdoor microcosms with and without antagonists (competitive and predatory invertebrates). Microcosms were treated repeatedly (three times) with thiacloprid at a concentration of 0.1, 1, or 10 µg/liter. In microcosms without antagonists, the abundance of Cx. pipiens larvae decreased moderately after the second and the third exposures to 10 µg/liter thiacloprid. In microcosms with antagonists, the abundance of Cx. pipiens larvae declined to approximately zero in the control group and the low concentration treatments during the five weeks of observation. By contrast, the abundance of Cx. pipiens larvae temporarily increased at 10 µg/liter thiacloprid after the second and third contamination. We explained this positive effect on the development of Cx. pipiens because of the decrease in competition due to the elimination of sensitive antagonists combined with the high recovery potential of Cx. pipiens. Based on these results, natural antagonists must be supported for the sustainable control of mosquitoes.     

Salt-marsh Crustaceans, Gammarus duebeni and Palaemonetes varians as Predators of Mosquito Larvae and Their Reaction to Bacillus thuringiensis subsp. israelensis

The predatory potential of two onmnivorous crustaceans, Gammarus duebeni and Palaemonetes varians, has been examined to investigate their effect as mosquito larval predators. Mature gammarids ate 4-8 Aedes detritus larvae/24 h and palaemonids, 22-30/1 h, often killing larvae when not hungry. The crustaceans were exposed to Bacillus thuringiensis subsp. israelensis (Bti) and to mosquito larvae killed by Bti with no adverse effects, endorsing the safety of this microbial pesticide. Crustacean faecal pellets, collected post-feeding and placed in fresh marsh water, were toxic to mosquito larvae the following day. After placing the crustaceans in fresh salt-marsh water for 6 days, the fresh faecal pellets failed to kill mosquitoes apart from pellets obtained from crustaceans originally fed on the highest concentration of Bti, where there was a 50% kill after 3 days of incubation. Mosquito larvae on salt-marshes are not easy to control with ecologically undesirable toxic chemicals. Encouraging the breeding of predators such as crustaceans, or even their release, could prove to be a useful method of mosquito control to supplement the periodic inundative use of the ecologically acceptable Bti.     

Chilodonella uncinata is Not Pathogenic to Culex pipiens Mosquitoes

Chilodonella uncinata was previously reported to be pathogenic to Culex and other mosquitoes based on the observation of high densities of ciliates inside of dead mosquito larvae. Here, we tested whether co‐culturing Cx. pipiens with Ch. uncinata resulted in decreased survival of the mosquito larvae. We find no evidence that Ch. uncinata is pathogenic to Cx. pipiens. We suggest that Ch. uncinata may instead feed on already deceased larvae.     

Oviposition site selection by mosquitoes is affected by cues from conspecific larvae and anuran tadpoles

Abstract The oviposition site that a female mosquito selects will influence the fitness of her larvae. We conducted a series of artificial pond experiments to compare the oviposition responses of two species of mosquitoes with the presence of tadpoles, conspecifics and chemical cues from these organisms. The two mosquito species differ markedly in larval ecology. The larvae of one species, Culex quinquefasciatus, co‐occur with numerous freshwater organisms, including tadpoles of Linmodynastes peronii (the striped marsh frog). Larvae of the other mosquito, Ochlerotatus australis, inhabit small brackish rock ponds where the main potential competitors are tadpoles of Crinia signifera (the common eastern froglet). In field trials, females of both mosquito species oviposited significantly more often in water that contained (or had previously contained) conspecific larvae. However, these superficially similar responses were mediated via different pathways: fungicide abolished the response by C. quinquefasciatus but not by O. australis. The two mosquito species also responded differently to cues associated with syntopic tadpoles. The presence of tadpoles did not influence oviposition by C. quinquefasciatus, but O. australis oviposited less often if tadpoles were present. These interspecific differences in oviposition behaviour may be adaptive to differences in larval ecology: competition with tadpoles is likely to be more significant for O. australis than for C. quinquefasciatus. Our findings thus support the hypothesis that mosquitoes oviposit selectively to avoid potential anuran larval competitors.     

Nestedness patterns of container-dwelling mosquitoes: effects of larval habitat within variable terrestrial matrices

Distributions of mosquito larvae likely are a consequence of multiple factors, although two commonly studied factors (quality of the larval environment and the terrestrial matrix in which these habitats reside) have rarely and simultaneously been varied in the field to understand distributions of larvae. We monitored aquatic containers of two leaf detritus levels within a forest, prairie, and industrial habitat across five months to understand the temporal and spatial colonization of aquatic invertebrates in Northcentral Illinois, USA. Data were collected monthly on mosquito populations and the composition of other invertebrates colonizing containers. Overall, six species of mosquitoes colonized containers, with Culex restuans and Aedes triseriatus having the highest relative abundances. There were strong seasonal abundance patterns for these two mosquito species, with the dominant species changing over time in the forest habitat. The responses of other mosquito taxa were more variable, with abundances reflective of either the terrestrial matrix or larval habitat quality. High detritus containers supported the highest abundances of most species encountered, regardless of habitat. Non-mosquito taxa were less common numerically, but analyses suggested that some taxa, such as syrphid larvae, often co-occurred with mosquitoes. Nested subset analysis indicated communities were strongly nested, and that both habitat type and detritus level were important in explaining nested patterns of aquatic invertebrates. Our data show that both the larval habitat and the surrounding terrestrial matrix shape patterns of container mosquitoes, and that other container invertebrates vary in similar ways as mosquitoes. Handling editor: K. Martens     

Larval habitat for the avian malaria vector Culex quinquefasciatus (Diptera: Culicidae) in altered mid‐elevation mesic‐dry forests in Hawai'i

Effective management of avian malaria (Plasmodium relictum) in Hawai'i's endemic honeycreepers (Drepanidinae) requires the identification and subsequent reduction or treatment of larval habitat for the mosquito vector, Culex quinquefasciatus (Diptera: Culicidae). We conducted ground surveys, treehole surveys, and helicopter aerial surveys from 2001–2003 to identify all potential larval mosquito habitat within two 100+ ha mesic‐dry forest study sites in Hawai'i Volcanoes National Park, Hawai'i; ‘Ainahou Ranch and Mauna Loa Strip Road. At ‘Ainahou Ranch, anthropogenic sites (43%) were more likely to contain mosquitoes than naturally occurring (8%) sites. Larvae of Cx. quinquefasciatus were predominately found in anthropogenic sites while Aedes albopictus larvae occurred less frequently in both anthropogenic sites and naturally‐occurring sites. Additionally, moderate‐size (≈20–22,000 liters) anthropogenic potential larval habitat had >50% probability of mosquito presence compared to larger‐ and smaller‐volume habitat (<50%). Less than 20% of trees surveyed at ‘Ainahou Ranch had treeholes and few mosquito larvae were detected. Aerial surveys at ‘Ainahou Ranch detected 56% (95% CI: 42–68%) of the potential larval habitat identified in ground surveys. At Mauna Loa Strip Road, Cx. quinquefasciatus larvae were only found in the rock holes of small intermittent stream drainages that made up 20% (5 of 25) of the total potential larval habitat. The volume of the potential larval habitat did not influence the probability of mosquito occurrence at Mauna Loa Strip Road. Our results suggest that Cx. quinquefasciatus abundance, and subsequently avian malaria, may be controlled by larval habitat reduction in the mesic‐dry landscapes of Hawai'i where anthropogenic sources predominate.     

The mosquito and predatory insect fauna inhabiting fresh-water ponds,with particular reference to Culex annulirostris Skuse (Diptera: Culicidae)

Colonization and abundance of mosquitoes and other insects were studied during four trials in man-made ponds at Mildura, Victoria. In December and February, respectively, Culex australicus and Cx annulirostris colonized the ponds within 1 day of formation. Maximum larval densities were attained within 8 days and, thereafter, densities gradually declined. The ponds were also colonized initially by predatory Coleoptera and Hemiptera, then Odonata. In three of the four trials, a significant inverse relationship existed between mosquito and predator densities. From life tables compiled during one of these trials, a survival rate of Cx annulirostris, from egg hatch to eclosion, was estimated to be 11%. Predation was considered to be largely responsible for the low survival. The results are discussed in relation to control of Cx annulirostris in temporary and permanent grassy ponds.     

Effects of nutrition and density in Culex pipiens

Mosquito larvae face numerous biotic and abiotic challenges that affect their development and survivorship, as well as adult fitness. We conducted two experiments under semi‐natural conditions to evaluate the effects of intraspecific competition, nutrient limitation and sub‐lethal doses of malathion on individual life history traits in adult Culex pipiens (Diptera: Culicidae). In the first experiment, larvae of Cx. pipiens were reared at different intraspecific densities and exposed to sub‐lethal doses of malathion. In the second experiment, different intraspecific densities of Cx. pipiens larvae were reared under conditions of low or high larval nutrients, and subsequent adults were fed on either water or 10% sucrose solution. Malathion treatment had relatively minor effects compared with density, which had significant negative effects on development rate, survivorship to adulthood, body size (wing length) and longevity. As larval density increased, a sex ratio distortion in survivorship to adulthood emerged, in which a bias towards males was apparent. Nutrient‐rich larval environments alleviated, in part, the effects of increasing density and extended the lifespan of mosquitoes fed on water and 10% sucrose. Density‐dependent alterations in adult longevity attributable to the larval environment are complex and show contrasting results depending on interactions with other environmental factors. This study suggests that larval resource availability and competition influence Cx. pipiens population growth correlates and have lasting effects on traits that relate to a mosquito's ability to vector pathogens.     

The mosquito fauna of the western region of Spain with emphasis on ecological factors and the characterization of Culex pipiens forms

This study updates the diversity, distribution, and seasonal trends of mosquitoes in a western region of Spain, assesses ecological determinants of Culex pipiens s.l., and determines form composition of Cx. pipiens s.s. populations. A total of 1,495 mosquitoes of 16 species was collected during 2012–2013, of which Cx. pipiens s.l. and Cx. theileri were the most abundant. Five new records for An. maculipennis s.s., Orthopodomyia pulcripalpis, Aedes (Ochlerotatus) punctor, Cx. europaeus, and Cx. modestus were found for this region. Cx. pipiens density varied across weather and habitat patterns, correlating positively with high temperatures and with a preference for urbanized areas and rural areas within a proximity of ovine farms. Moreover, molecular identification by CQ11FL was performed in 467 Cx. pipiens s.s., detecting both pipiens (66%) and molestus (8.4%) forms coexisting in different habitats (urban, peri‐urban and rural) aboveground with a high degree of hybridization (25.7%). The abundance of Cx. pipiens in urban areas and farms, with the presence of hybrids, may increase their capacity to act as bridge vectors for the transmission of arboviral infections. These data will be helpful for further implementation of entomological programs focused on risk assessment for arboviruses or other mosquito‐borne pathogens.     

Characterisation of Culex quinquefasciatus (Diptera: Culicidae) larval habitats at ground level and temporal fluctuations of larval abundance in Córdoba, Argentina

The aims of this study were to characterise the ground-level larval habitats of the mosquito Culex quinquefasciatus, to determine the relationships between habitat characteristics and larval abundance and to examine seasonal larval-stage variations in Córdoba city. Every two weeks for two years, 15 larval habitats (natural and artificial water bodies, including shallow wells, drains, retention ponds, canals and ditches) were visited and sampled for larval mosquitoes. Data regarding the water depth, temperature and pH, permanence, the presence of aquatic vegetation and the density of collected mosquito larvae were recorded. Data on the average air temperatures and accumulated precipitation during the 15 days prior to each sampling date were also obtained. Cx. quinquefasciatus larvae were collected throughout the study period and were generally most abundant in the summer season. Generalised linear mixed models indicated the average air temperature and presence of dicotyledonous aquatic vegetation as variables that served as important predictors of larval densities. Additionally, permanent breeding sites supported high larval densities. In Córdoba city and possibly in other highly populated cities at the same latitude with the same environmental conditions, control programs should focus on permanent larval habitats with aquatic vegetation during the early spring, when the Cx. quinquefasciatus population begins to increase.     

Successional mosquito dynamics in surrogate treehole and ground‐container habitats in the northeastern United States: Where does Aedes albopictus fit in?

This study assessed the risk of larval displacement of the eastern treehole mosquito, Aedes triseriatus, and the northern house mosquito, Culex pipiens, by Aedes albopictus, the Asian tiger mosquito, during the establishment and successional stages of novel larval mosquito treehole and ground‐container habitats in the state of New Jersey, U.S.A. Culex pipiens and Culex restuans were the first mosquito species to colonize ground‐container habitats and were the dominant larval species throughout the study period, whereas Ae. albopictus was late to colonize ground habitats and accounted for less than 15% of weekly larval collections once established. Ae. albopictus had a much stronger community presence within treehole ovitraps; however, Ae. albopictus never reached the average larval densities of the expected primary colonizer, Ae. triseriatus. Throughout the study period, the weekly abundances of Ae. triseriatus and Ae. albopictus were positively correlated and there were no significant differences between the abundances of each species. The larval dominance of Ae. triseriatus appears to be enhanced by the presence of Toxorhynchites rutilus septentrionalis, a large predatory mosquito species. When Tx. rut. septentrionalis was present, mature larvae (3^rd–4^th instar) of Ae. albopictus were also present in only 16.7% of collections, whereas mature larvae of Ae. triseriatus were collected concurrently with Tx. rut. septentrionalis in 53.8% of collections. These data suggest that Ae. triseriatus is at a greater risk of displacement by Ae. albopictus than are Cx. pipiens and Cx. restuans.     

Unraveling microbe-mediated interactions between mosquito larvae in a laboratory microcosm

Interspecies interactions have important impacts on communities and when multiple trophic levels are involved, effects can be complex and indirect. For mosquitoes, interactions experienced as larvae affect adult attributes such as survivorship, reproductive output, and longevity, factors that can affect their ability to vector disease. We examined how larvae of two ecologically distinct mosquito species, Aedes japonicus japonicus and Culex quinquefasciatus, interact at different temperatures (17 and 27 °C) and at different relative densities. We also quantified abundances of bacteria and protozoan flagellates to uncover how changes in the microbial community affect the outcome of the two mosquitoes’ interaction. At 17 °C, survival and size of both mosquito species were not affected by the other’s presence. Cx. quinquefasciatus was strongly affected by intraspecific, but not interspecific, competition at both temperatures. At 27 °C, Ae. j. japonicus larvae experienced 100 % mortality in treatments by themselves and treatments where Cx. quinquefasciatus was abundant, surviving only in the presence of low densities of Cx. quinquefasciatus. Both the total bacteria count and counts of a protozoan flagellate identified as Spumella spp. decreased with increasing numbers of Cx. quinquefasciatus. We postulate that at 27 °C, the survival of Ae. j. japonicus depends on the interaction between Cx. quinquefasciatus and the microbial community. This study demonstrates that one mosquito species may alter the microbial community in ways that indirectly influence another mosquito species’ larval survival, and by extension adult abundance and potential disease transmission.     

Effects of flash flooding on mosquito and community dynamics in experimental pools

Pulsed disturbances of larval mosquito sites are likely to have a direct negative effect on mosquitoes but may also have indirect effects due to the alteration of community structure. These altered communities may become attractive to gravid mosquitoes searching for oviposition sites when the disturbances decrease the abundance of mosquito antagonists such as competitors, which often results in an increase in mosquito food resources. However, flash flood disturbances in intermittent riverbeds can also remove mosquito food resources such as algae, so that the net effect of flash floods could be either to increase or decrease mosquito abundance. We conducted an outdoor mesocosm experiment to assess the effects of flash floods on mosquito oviposition habitat selection and larval abundance during the post‐disturbance period of community recovery. Mesocosms were artificially flooded. Mosquito oviposition, immature abundance, invertebrate species diversity, chlorophyll a, and abiotic parameters were monitored. Our results showed that the flash flood negatively affected phytoplankton and zooplankton, leading to a decrease of mosquito oviposition in flooded mesocosms compared to non‐flooded mesocosms. More broadly, this study indicates how disturbances influence mosquito oviposition habitat selection due to the loss of food resources in ephemeral pools, and it highlights the importance of considering the effects of disturbances in management, habitat restoration, and biodiversity conservation in temporary aquatic habitats.     

Can adults of the New Zealand mosquito Culex pervigilans (Bergorth) detect the presence of a key predator in larval habitats?

The influence of predators on mosquito populations may be direct through predation or indirect through sub‐lethal responses of adult mosquitoes in life history traits such as oviposition behavior. In New Zealand, the backswimmer, Anisops wakefieldi, is a common predator of mosquito larvae found in temporary and permanent water bodies. We predicted that the New Zealand native mosquito, Culex pervigilans, whose larvae are vulnerable to predation of Anisops, would likely avoid the containers with the presence of Anisops or its kairomone. We established temporary water containers without predators, free‐roaming predators, caged predators (which were unable to eat mosquitoes), or containers from which predators were removed immediately prior to the experiment (these containers would have remnant kairomones from the predators). Each treatment with Anisops had predator densities of one, three, or nine Anisops. Contrary to our predictions, when choosing oviposition habitats, Cx. pervigilans appeared to ignore the presence of free‐roaming Anisops, caged Anisops, and water with Anisops kairomone. We thus observed no significant differences between the numbers of egg rafts laid by Cx. pervigilans in the different predator treatments nor were the number of egg rafts significantly affected by the density of predators. Rather than the presence of predators, environmental factors including temperature, humidity, and pressure were significantly correlated with mosquito oviposition. These mosquitoes appeared to either ignore the presence of the predator, had no ability to detect the presence of predators, or perhaps the cues from Anisops predators were not sufficiently strong enough to alarm these mosquitoes. We argue that the mosquito has not evolved the ability to detect the presence of these predators while ovipositing.