Culex erythrothorax (Diptera: Culicidae): Activity periods, insecticide susceptibility and control in California (USA).

The mosquito Culex erythrothorax Dyar is a West Nile virus (WNV) vector that breeds in wetlands with emergent vegetation. Urbanization and recreational activities near wetlands place humans, birds and mosquitoes in close proximity, increasing the risk of WNV transmission. Adult Cx. erythrothorax abundance peaked in a wetland bordering the San Francisco Bay of California (USA) during the first 3 hours after sunset (5527 ± 4070 mosquitoes / trap night) while peak adult Culex tarsalis Coquillett abundance occurred during the subsequent 3 h period (83 ± 30 Cx. tarsalis). When insecticide resistance was assessed using bottle bioassay, Cx. erythrothorax was highly sensitive to permethrin, naled, and etofenprox insecticides compared to a strain of Culex pipiens that is susceptible to insecticides (LC50 = 0.35, 0.71, and 4.1 µg/bottle, respectively). The Cx. erythrothorax were 2.8-fold more resistant to resmethrin, however, the LC50 value was low (0.68 µg/bottle). Piperonyl butoxide increased the toxicity of permethrin (0.5 µg/bottle) and reduced knock down time, but a higher permethrin concentration (2.0 µg/bottle) did not have similar effects. Bulk mixed-function oxidase, alpha-esterase, or beta-esterase activities in mosquito homogenates were higher in Cx. erythrothorax relative to the Cx. pipiens susceptible strain. There was no difference in the activity of glutathione S-transferase between the two mosquito species and insensitive acetylcholine esterase was not detected. Larvicides that were applied to the site had limited impact on reducing mosquito abundance. Subsequent removal of emergent vegetation in concert with larvicide applications and reduced daily environmental temperature substantially reduced mosquito abundance. To control Cx. erythrothorax in wetlands, land managers should consider vegetation removal so that larvicide can efficiently enter the water. Vector control agencies may more successfully control adult viremic Cx. erythrothorax that enter nearby neighborhoods by applying adulticides during the 3 h that follow sunset.

Evidence of transovarial transmission of Chikungunya and Dengue viruses in field-caught mosquitoes in Kenya.

Arboviruses are among the most important emerging pathogens due to their increasing public health impact. In Kenya, continued population growth and associated urbanization are conducive to vector spread in both urban and rural environments, yet mechanisms of viral amplification in vector populations is often overlooked when assessing risks for outbreaks. Thus, the characterization of local arbovirus circulation in mosquito populations is imperative to better inform risk assessments and vector control practices. Aedes species mosquitoes were captured at varying stages of their life cycle during different seasons between January 2014 and May 2016 at four distinct sites in Kenya, and tested for chikungunya (CHIKV), dengue (DENV) and Zika (ZIKV) viruses by RT-PCR. CHIKV was detected in 45 (5.9%) and DENV in 3 (0.4%) mosquito pools. No ZIKV was detected. Significant regional variation in prevalence was observed, with greater frequency of CHIKV on the coast. DENV was detected exclusively on the coast. Both viruses were detected in immature mosquitoes of both sexes, providing evidence of transovarial transmission of these arboviruses in local mosquitoes. This phenomenon may be driving underlying viral maintenance that may largely contribute to periodic re-emergence among humans in Kenya.

Ovipositional and ovicidal effects of the microbial agent Bacillus thuringiensis israelensis on Culex quinquefasciatus say (Diptera: Culicidae).

The magnitude of oviposition as well as the size, shape and the number of eggs per of egg rafts egg raft were determined after gravid Culex quinquefasciatus Say oviposited on water treated with water dispersible granules (WDG) of Bacillus thuringiensis ssp. israelensis (Bti) and on untreated water. The mean number of eggs/raft was lower in the treated than in the untreated water. Bti concentrations from 0.5 to 2.0mg/L affected the shape of egg rafts and number of eggs in each raft. As the concentration of Bti increased from 0.5 to 2.0 mg/L the shape of egg rafts became more irregular with fewer eggs in each raft. Exposure to Bti at 2- and 26-h reduced the hatching rates, and fewer eggs hatched at 26-h of exposure to Bti. As the concentration of Bti WDG increased from 0.5 to 2.0 mg/L, the hatching rate decreased. Eggs exposed for 2-h to 2.0mg/ L Bti had a hatch of 30% after 24 h, the rate increasing to 57% after 72 h. In contrast, in 26-h exposed eggs to 2.0 mg/L Bti, the hatching rate after 24 h was only 12% and this rate increased to 39% after 72 h. In larvae from eggs exposed for 2 h, the survival rate was 40% at 2.0 mg/L Bti and 87% in untreated controls. In contrast, the survival rates of larvae from 26-h exposed eggs was 91% in controls while it was 30% at 2.0 mg/L Bti. As the concentration of Bti increased from 0.5 to 2.0 mg/ 1 the survival rates of larvae decreased. The combined effects of reductions of egg rafts, low number of eggs per egg raft, and reduced hatching and survival rates could have significant cumulative effects on the yield of adult mosquitoes, and this could result in a greater control potential of this microbial agent.

Non-larvicidal effects of Bacillus thuringiensis israelensis and Bacillus sphaericus on oviposition and adult mortality of Culex quinquefasciatus Say (Diptera: Culicidae).

Two microbial mosquito larvicides, Bacillus thuringiensis ssp. israelensis (Bti) and Bacillus sphaericus (Bsph), have been shown to be highly effective in controlling mosquito larvae and have been used in larvicidal programs for many years. In exploring other modes of action of these agents, we studied the ovipositional response of Bsph susceptible and resistant Culex quinquefasciatus to aqueous suspensions of Bti and Bsph water dispersible granules (WDG). We quantified the level of mortality of adult mosquitoes caused by exposure to Bti and Bsph suspensions during oviposition. Significantly lower numbers of egg rafts were laid and collected from the treatments than the control regimen. There was an inverse relationship between Bacillus product concentrations and oviposition. As the concentration of Bti or Bsph increased from 0.0 to 2.0 mg/L, treated waters received progressively fewer egg rafts. In addition to the negative effects of Bacillus on oviposition, both male and female adult mosquitoes suffered high mortality on landing and imbibing on Bti and Bsph suspensions, the extent of mortality directly proportional to concentration. These two microbial agents used solely as mosquito larvicides thus have the additional benefits of reducing mosquito oviposition and killing adult mosquitoes, especially gravid females that come in contact with the treated water either for oviposition or drinking. Reducing the number of gravid females may also result in reduced transmission rates of pathogens. The combined effects of reduced oviposition and adult mortality could result in higher control potential of these microbial agents.

Mosquito larvicidal activity of aqueous extracts of long pepper (Piper retrofractum vahl) from Thailand.

Aqueous extracts of nine medicinal plants were bioassayed against larvae of Culex quinquefasciatus Say and Aedes aegypt (L.). Among these plants, the long pepper, Piper retrofractum Vahl (Piperaceae), showed the highest level of activity against mosquito larvae. To gain more information on larvicidal activity of P. retrofractum, fresh fruits of this plant were extracted in water and the extracts made into powder and bioassayed against 3rd and 4th instar larvae of Cx. quinquefasciatus and Ae. aegypti in the laboratory. Extracts of unripe (001/3) and ripe (002/3 and 001/4) fruits showed different levels of activity against Cx. quinquefasciatus larvae. Extracts 001/3 and 002/3 were equi-toxic to a Bacillus sphaericus resistant and susceptible strains, both from Thailand. The ripe fruit extract 002/3 was somewhat more active against Ae. aegypti than Cx. quinquefasciatus. Another ripe fruit extract (001/4) was much more toxic to both mosquito species. Diluted solutions of the solid extract (002/3) in distilled water lost their larvicidal activity upon aging. Loss of activity at 25 degrees C was greater than that stored at 4 degrees C, and greater in water than in acetone solution.

Laboratory and simulated field evaluation of a new recombinant of Bacillus thuringiensis ssp. israelensis and Bacillus sphaericus against Culex mosquito larvae (Diptera: Culicidae).

In the laboratory, three microbial mosquito larvicidal products consisting of Bacillus thuringiensis ssp. israelensis de Barjac (Bti), Bacillus sphaericus (Neide) (Bsph) (strain 2362), and the University of California Riverside (UCR) recombinant (producing toxins of both Bacillus sphaericus and Bacillus thuringiensis ssp. israelensis) were bioassayed against larvae of Culex quinequefasciatus Say (susceptible and resistant to Bsph 2362), and Aedes aegypti (L.). Bti proved highly effective against Cx. Quinequefasciatus susceptible and resistant strains, with LC50 values of 0.009 and 0.011 ppm and LC90 values of 0.057 and 0.026 ppm for Bsph-susceptible and -resistant strains, respectively. Bti was also highly active against Ae. eagypti with LC50 and LC90 values of 0.014 and 0.055 ppm, respectively. The UCR recombinant was equally active against both Bsph-susceptible and -resistant strains of Cx. Quinquefasciatus; LC50 values were 0.005 and 0.009 and LC90 values were 0.030 and 0.043 ppm, respectively. Bti and the UCR recombinant essentially showed similar activity against Bsph-susceptible and -resistant strains. UCR recombinant showed high toxicity against Ae. eagypti with LC50 and LC90 values of 0.023 and 0.064 ppm, respectively. Bsph was highly active against susceptible strain of Cx. quinequefasciatus with LC50 and LC9o values of 0.006 and 0.024 ppm, respectively. Bsph exhibited little toxicity against Ae. eagypti larvae and also no toxicity to Bsph resistance. In the field, we evaluated four experimental corn grit formulations of Bti (VBC 60021), Bsph (VBC 60022), UCR recombinants VBC 60023 (7.89%), and VBC 60024 (1.87%) in simulated field (microcosms) against Bsph-susceptible Culex mosquitoes. Bti and low-concentrate UCR recombinant showed similar initial activity as well as persistence. Both materials provided high-to-moderate level of control for 2-7 d posttreatment at low treatment rates. At low dosages, residual activity of Bti and UCR recombinant lasted for <7 d. Bsph and high-concentrate UCR recombinant (VBC 60023), however, were more effective against natural populations of Cullex and achieved longer control (7-21 d) than the other two materials.

Susceptibility profile of Culex quinquefasciatus (Diptera: Culicidae) to Bacillus sphaericus on selection with rotation and mixture of B. sphaericus and B. thuringiensis israelensis.

Several strains of Bacillus sphaericus (Neide) (Bsph), a spore-forming bacterium that produces potent mosquitocidal toxins, have been widely used against mosquito larvae globally. In some locations in the field and in the laboratory, moderate to high levels of resistance in Culex pipiens complex mosquito larvae to Bsph have been reported. Practical strategies for dealing with the resistance problem and more importantly to delay or possibly prevent development of resistance to Bsph in the first place are urgently needed. From published reports, we deduced that Bsph-resistant mosquito larvae are completely susceptible to Bacillus thuringiensis ssp. isreaelensis (Bti) and that there is no appearance of cross-resistance to Bti. On the basis of this information, we hypothesized that Bti offers a good potential for the management of resistance to Bsph, especially when it comes to delaying or preventing emergence of resistance. We conducted laboratory studies to see whether rotation of Bsph and Bti or mixture of both delays or prevents resistance development in Bsph-susceptible California Cx. quinquefasciatus. Development of resistance using Bsph alone increased after F15 and fluctuated slightly on further selections. Rotation of Bsph and Bti resulted in much higher level and rapid emergence of resistance to Bsph. However, selection with mixtures of Bti and Bsph for 36 generations showed no emergence of resistance. On the basis of this information, we believe mixtures have a good potential in delaying or preventing Bsph resistance.

Strategies for the management of resistance in mosquitoes to the microbial control agent Bacillus sphaericus.

Bacillus sphaericus (B.spi) strain 2362 has been recognized as a promising mosquito larvicide, and various preparations of this strain have been tested and used in mosquito control programs worldwide. This control agent has advantages of high efficacy, specificity, persistence, and environmental safety. However, resistance in Culex pipiens complex mosquitoes to Bsph has occurred in both laboratory and field populations, necessitating development of resistance management strategies. Studies were initiated aiming at reversing previously established Bsph resistance in a laboratory colony of Culex quinque fasciatus Say by selections with Bti alone, Bti and Bsph in rotation, or mixture. Partial restoration of susceptibility to Bsph was achieved by selection of resistant colony for 10 generations with Bti alone at LC80). After this colony was switched back to Bsph selection for 20 generations, resistance to Bsph partially increased to a stable level. Selections of Basph-resistant colonies with Bti and Bsph in rotation or mixture resulted in steady decline of resistance over 30 generations, with rapid decline in resistance noted in the initial 10-15 generations. It is interesting to note that selections with Bti and Bsph in rotation increased susceptibility to Bti in Bsph-resistant colony. It is promising that selection with Bti alone, Bsph and Bti in rotation, or mixture have a potential for developing practical strategies to overcome acquired resistance to Bsph in Cx. quinquefasciatus populations.