Microbial Larvicide Research Articles

Effect of Repeated Application of Microbial Larvicides on Malaria Transmission in Central Côte d'Ivoire

The effect of repeated applications of Bacillus thuringiensis var israeliensis (Bti) and B. sphaericus (Bs) on different entomologic parameters of malaria transmission was investigated in a village in central Côte d'Ivoire. The study area was monitored for potential mosquito breeding sites over a 7-month period. Microbial larvicides were applied once every 3 wk; first Bti (0.8 mg/liter), followed by Bs (10 mg/liter) 3-4 days later. Adult mosquitoes were collected inside and outside sentinel houses in 4 cross-sectional surveys using human landing catch. Repeated applications of Bti and Bs showed a decline in the biting rate of both Anopheles funestus and Anopheles gambiae. Moreover, the entomologic inoculation rate of An. funestus was significantly reduced (from 328 to 142, P = 0.005), whereas that of An. gambiae remained stable. In conclusion, microbial larvicides might play a role in an integrated approach for malaria control.

Evaluation of Three Microbial Formulations Against Culex pipiens pipiens Larvae in Irrigation Fields in Wroclaw, Poland

Two evaluations of microbial mosquito larvicides were conducted in irrigation fields located along the Odra River near the city of Wroclaw, Poland. In the first evaluation, 3 dosages each of Bacillus thuringiensis israelensis (Bti)--VectoBac water dispersible granule (WDG) and VectoBac aqueous suspension formulation (12AS)--and Bacillus sphaericus (Bs)--VectoLex WDG--were tested against larvae of Culex pipiens pipiens. There was little difference between formulations and rates of application between day 2 and 7 following the treatment. VectoBac WDG at 300 g/ha resulted in 100% mortality of Cx. p. pipiens. Between 0.5 and 1 liter of VectoBac 12AS caused sufficient mortality for up to 7 days. The effective shortterm application rate of VectoLex WDG was found to be 400 g/ha. The above doses can be recommended for up to 7-day control in this habitat. In the second evaluation, using VectoBac WDG at 800 g/ha, the residual efficacies of 2 ground application methods, a knapsack sprayer configured for large droplet application and a motorized backpack blower configured for small droplet application, were compared. The results indicated no significant difference between the 2 application methods, but a significant long-term effect of VectoBac WDG was observed at the rate of 800 g/ha.

Microbial larvicides for malaria control in The Gambia

BackgroundMosquito larval control may prove to be an effective tool for incorporating into integrated vector management (IVM) strategies for reducing malaria transmission. Here the efficacy of microbial larvicides against Anopheles gambiae s.l. was tested in preparation for a large-scale larviciding programme in The Gambia.MethodsThe impact of water-dispersible (WDG) and corn granule (CG) formulations of commercial Bacillus sphaericus strain 2362 (Bs; VectoLex®) and Bacillus thuringiensis var.israelensis strain AM65-52 (Bti; VectoBac®) on larval development were tested under laboratory and field conditions to (1) identify the susceptibility of local vectors, (2) evaluate the residual effect and re-treatment intervals, (3) test the effectiveness of the microbials under operational application conditions and (4) develop a method for large-scale application.ResultsThe major malaria vectors were highly susceptible to both microbials. The lethal concentration (LC) to kill 95% of third instar larvae of Anopheles gambiae s.s. after 24 hours was 0.023 mg/l (14.9 BsITU/l) for Bs WDG and 0.132 mg/l (396 ITU/l) for Bti WDG. In general Bs had little residual effect under field conditions even when the application rate was 200 times greater than the LC95. However, there was a residual effect up to 10 days in standardized field tests implemented during the dry season. Both microbials achieved 100% mortality of larvae 24–48 hours post-application but late instar larvae were detected 4 days after treatment. Pupae development was reduced by 94% (95% Confidence Interval = 90.8–97.5%) at weekly re-treatment intervals. Field tests showed that Bs had no residual activity against anopheline larvae. Both microbials provided complete protection when applied weekly. The basic training of personnel in identification of habitats, calibration of application equipment and active larviciding proved to be successful and achieved full coverage and control of mosquito larvae for three months under fully operational conditions.ConclusionEnvironmentally safe microbial larvicides can significantly reduce larval abundance in the natural habitats of The Gambia and could be a useful tool for inclusion in an IVM programme. The costs of the intervention in this setting could be reduced with formulations that provide a greater residual effect.

Transposon-mediated resistance to Bacillus sphaericus in a field-evolved population of Culex pipiens (Diptera: Culicidae)

The binary toxin is the major active component of Bacillus sphaericus, a microbial larvicide used for controlling some vector mosquito-borne diseases. B. sphaericus resistance has been reported in many part of the world, leading to a growing concern for the usefulness of this environmental friendly insecticide. Here we characterize a novel mechanism of resistance to the binary toxin in a natural population of the West Nile virus vector, Culex pipiens. We show that the insertion of a transposable element-like DNA into the coding sequence of the midgut toxin receptor induces a new mRNA splicing event, unmasking cryptic donor and acceptor sites located in the host gene. The creation of the new intron causes the expression of an altered membrane protein, which is incapable of interacting with the toxin, thus providing the host mosquito with an advantageous phenotype. As a large portion of insect genomes is composed of transposable elements or transposable elements-related sequences, this new mechanism may be of general importance to appreciate their significance as potent agents for insect resistance to the microbial insecticides.

Sublethal Effects of Mosquito Larvicides on Swimming Performance of Larvivorous Fish Melanotaenia duboulayi (Atheriniformes: Melanotaeniidae)

Laboratory studies were conducted to determine the sublethal effects of exposure to selected larvicides on the critical swimming speed (Ucrit) of crimson-spotted rainbowfish, Melanotaenia duboulayi (Castlenau). This native fish is common throughout southeastern Queensland, and it is increasingly being distributed as a biological control agent of mosquitoes. The selected larvicides included, two organophosphate (OP) compounds (temephos and pirimiphos-methyl), two microbial larvicides (Bacillus thuringiensis spp. israelensis [Bti] de Barjac and Bacillus sphaericus [Bs] Neide), and an insect growth regulator (IGR) (s-methoprene). Exposure to the OP temephos at 10 times the effective field concentration (EFC; 0.33 mg/liter), and OP pirimiphos-methyl at the EFC (0.50 mg/liter), resulted in a significant reduction in the Ucrit of M. duboulayi under controlled conditions. Conversely, exposure to the microbial (Bti and Bs) and IGR (s-methoprene) larvicides at 10 times the EFC had no effect on the Ucrit of M. duboulayi. Accordingly, these products are suitable for integrated pest management programs in Australia.

Suppression of exposure to malaria vectors by an order of magnitude using microbial larvicides in rural Kenya

To determine the contribution larviciding could make to reduce the burden of malaria, by conducting a trial of microbial larvicides in a 4.5 km2 area in and around a large village in rural western Kenya. The abundance of immature and adult mosquitoes was monitored for 12 months under baseline conditions. Then microbial larval control was implemented for 28 months. After the intervention, the abundance of immature and adult mosquitoes was monitored for a further 12 months. Of the 419 mosquito larval habitats identified, 336 (80%) originated from human activities. Application of Bacillus thuringiensis var. israelensis and Bacillus sphaericus larvicides reduced the proportion of aquatic habitats containing Anopheles larvae from 51% during non-intervention periods to 7% during the intervention. The occurrence of late instar Anopheles in habitats was reduced from 39% and 33% in pre-intervention and post-intervention periods to 0.6% during intervention. Overall, larviciding reduced Anopheles larval density by 95% and human exposure to bites from adults by 92%. The estimated cost of providing this protection to the human population in the study area was less than US$ 0.90/person/year. Appropriately applied microbial larvicides can substantially and cost-effectively reduce human exposure to malaria in rural sub-Saharan Africa.

Microbial AgentsBacillus thuringiensisssp.israelensisandBacillus sphaericusSuppress Eutrophication, Enhance Water Quality, and Control Mosquitoes in Microcosms

Bacillus thuringiensis ssp. israelensis de Barjac ( Bti ) and Bacillus sphaericus Neide are promising microbial mosquito larvicides currently employed in mosquito control programs. During the screening and evaluation of new formulations of these control agents, it was noted that productivity of some algal species was noticeably suppressed, and that certain water quality parameters were enhanced by the suppression of primary productivity with these agents. Quantitative studies on these aspects were conducted in microcosms where Culex mosquitoes bred naturally. Treatments with VectoBac G ( Bti ) at the high dosage of 48.1 kg/ha, B. sphaericus water dispersible granules at 3.1 kg/ha, and Bti water dispersible granules at 0.6 kg/ha yielded good control of immature mosquito populations. Two species of unicellular algae, Closterium sp. and Chlorella sp., existed in both the controls and treatments. Growth of these algae was suppressed by the microbial agents applied. Suppressed algal productivity and photosynthesis resulted in lower water turbidity and oxygen concentrations in the treatments than in the controls, especially during the hot season. Water in treatments was discernibly clearer than in the controls, which was further borne out by differences in the optical density readings.

Effects of Bacillus thuringiensis israelensis (BTI) and Methoprene on Nontarget Macroinvertebrates in Minnesota Wetlands

We studied the effects of the mosquito larvicides methoprene and Bacillus thuringiensis israelensis (Bti) on the benthic macroinvertebrate communities of 27 wetland ecosystems in Wright County, Minnesota. These larvicides are generally considered safe for nontarget species. After 3 yr of preliminary investigations, including 2 yr of intensive sampling, larvicide treatments were applied during 1991–1993. Nine of the wetlands were experimentally treated with methoprene, which disrupts insect development; an additional set of nine wetlands were treated with Bti, a microbial larvicide; and nine wetlands were left untreated to serve as a control treatment. In general, insecticide treatment had minimal effects on nontarget groups during the first treatment year. However, during 1992, highly significant reductions due to both methoprene and Bti were observed in several insect groups. Predatory insects were reduced on methoprene-treated sites but not Bti-treated sites in 1992. In 1993, treatment with both larvicides resulted in wetland communities that were depauperate in most insects. Although effects were observed broadly across insect taxa, Diptera, which comprised 79% of the insects, were affected most strongly, especially the dipteran suborder Nematocera, which included 71% of total insects and was dominated by Chironomidae. Minimal effects on noninsect macroinvertebrates were observed. Bti- and methoprene-treated sites also showed a reduction in richness of insect genera and an increased tendency to be dominated by one or a few genera. Pretreatment data, collected under drought conditions from the same wetlands, showed that the benthic macroinvertebrate fauna was dominated by mollusks during the drought but became increasingly dominated by insects during the wetter years. On the treated sites, insects remained at low density, very similar to the drought conditions, but the noninsect macroinvertebrates declined on treated sites in the same pattern as on control sites. Both indirect effects and direct toxicity likely contributed to the observed differences. Bti is likely to be directly toxic only to nematoceran Diptera; thus effects of Bti on other insect groups may have resulted from disruption of the invertebrate food web. Methoprene is more broadly toxic; thus observed methoprene effects on nonnematoceran groups may have been due to either direct toxicity or food web effects, or both. The 2–3 yr lag time in response of nontarget insects to larvicide treatment demonstrates the need for long-term studies in wetland ecosystems, and the need to reconsider the conclusions based on previous short-term studies that these larvicides are environmentally safe.

EFFECTS OFBACILLUS THURINGIENSIS ISRAELENSIS(BTI) AND METHOPRENE ON NONTARGET MACROINVERTEBRATES IN MINNESOTA WETLANDS

We studied the effects of the mosquito larvicides methoprene and Bacillus thuringiensis israelensis (Bti) on the benthic macroinvertebrate communities of 27 wetland ecosystems in Wright County, Minnesota. These larvicides are generally considered safe for nontarget species. After 3 yr of preliminary investigations, including 2 yr of intensive sampling, larvicide treatments were applied during 1991–1993. Nine of the wetlands were experimentally treated with methoprene, which disrupts insect development; an additional set of nine wetlands were treated with Bti, a microbial larvicide; and nine wetlands were left untreated to serve as a control treatment. In general, insecticide treatment had minimal effects on nontarget groups during the first treatment year. However, during 1992, highly significant reductions due to both methoprene and Bti were observed in several insect groups. Predatory insects were reduced on methoprene-treated sites but not Bti-treated sites in 1992. In 1993, treatment with both larvicides resulted in wetland communities that were depauperate in most insects. Although effects were observed broadly across insect taxa, Diptera, which comprised 79% of the insects, were affected most strongly, especially the dipteran suborder Nematocera, which included 71% of total insects and was dominated by Chironomidae. Minimal effects on noninsect macroinvertebrates were observed. Bti- and methoprene-treated sites also showed a reduction in richness of insect genera and an increased tendency to be dominated by one or a few genera. Pretreatment data, collected under drought conditions from the same wetlands, showed that the benthic macroinvertebrate fauna was dominated by mollusks during the drought but became increasingly dominated by insects during the wetter years. On the treated sites, insects remained at low density, very similar to the drought conditions, but the noninsect macroinvertebrates declined on treated sites in the same pattern as on control sites. Both indirect effects and direct toxicity likely contributed to the observed differences. Bti is likely to be directly toxic only to nematoceran Diptera; thus effects of Bti on other insect groups may have resulted from disruption of the invertebrate food web. Methoprene is more broadly toxic; thus observed methoprene effects on nonnematoceran groups may have been due to either direct toxicity or food web effects, or both. The 2–3 yr lag time in response of nontarget insects to larvicide treatment demonstrates the need for long-term studies in wetland ecosystems, and the need to reconsider the conclusions based on previous short-term studies that these larvicides are environmentally safe.

Organic Matter Processing by Larval Black Flies in a Temperate Woodland Stream

We studied the role of a dense aggregation of filter-feeding black fly larvae in organic matter processing in a north temperate woodland stream. Black fly guts contained algae. individual bacterial cells, bacterial clusters, and detritus, but bacterial clusters and especially detritus were a much larger proportion of gut material. Particle size distribution and size variance within the largest size fraction of fine particulate organic matter (FPOM) studied (>100 μm) decreased downstream of black flies, due to the high proportion of uniform-sized fecal pellets in this size category. Ash free dry mass (AFDM) of FPOM was 28% higher downstream of the aggregation than upstream. even though black flies were actively feeding. As a control for factors other than black flies that might alter FPOM mass, we killed the black fly larvae with a highly specific microbial larvicide (Bacillus thuringiensis var. israelensis or Bti), then remeasured FPOM mass at the same stations. Following Bti treatment, FPOM mass was only about 5.6% higher downstream of the aggregation, an increase that was likely because FPOM flocculates from dissolved organic matter (DOM) due to turbulence. These data suggested that black flies were feeding from another pool of organic matter in addition to suspended FPOM, which we hypothesized to be DOM. Laboratory experiments using filtered (0.2 μm) stream water demonstrated that black flies ingested 14 C-labeled DOM. Our results indicate that the quantitative effects of black flies on FPOM in streams are linked to DOM ingestion. We suggest that a more complete examination of these processes is needed for a comprehensive understanding of organic matter processing in streams.

Comparative Ingestion Rates of Culex quinquefasciatus (Diptera: Culicidae) Susceptible and Resistant to Bacillus sphaericus

Larvae of laboratory and field-collected strains of Culex quinquefasciatus susceptible and resistant (37-and 31-fold, upon selection) to Bacillus sphaericus strain 2362 were studied for their ingestion rates of this pathogen and other particulate materials. Larvae of the resistant strains (laboratory and field-collected) ingested B. sphaericus preparation at significantly lower rates than did larvae of the susceptible strains. Larvae of the resistant strains also ingested B. sphaericus preparation at significantly lower rates than yeast (nutritious), while larvae of the susceptible strains ingested B. sphaericus preparation at the same or higher rates than yeast. The reduced ingestion rates on B. sphaericus in resistant strains could possibly serve as a behavioristic mechanism of resistance against this microbial larvicide.

Control of simuliid blackflies and mosquitoes withBacillus thuringiensis subsp.israelensis

Bacillus thuringiensis subsp.israelensis produces a δ-endotoxin which is insecticidal against larvae of mosquitoes and blackflies. The toxin particles introduced into the water at concentrations of a few ng/ml kill rapidly after uptake by target larvae. Breakdown of the function of the gut epithelium is the primary cause for the lethal action of the toxin. The molecular mode of action is not yet understood in its details. However, it appears that the toxin is cytolytic and acts by disruption of membrane permeability systems. Field efficacy ofB. thuringiensis subsp.israelensis has been demonstrated in many countries. Most impressive results were obtained in West Africa where this microbial larvicide has been included in a large field programme to control blackflies, the vector of river blindness.