Fate of Bacillus sphaericus and Bacillus thuringiensis serovar israelensis in the aquatic environment.

Bacillus sphaericus spores were suspended in bottles of filtered (0.45 microns) freshwater and seawater under various conditions of temperature, pH and salinity. Heat resistant culturable counts (spores) slowly decreased with time. Spores suspended in dialysis bags submerged in a freshwater pond or in flowing seawater underwent a more rapid drop in heat resistant spore counts than did spores held in bottles. Thus, laboratory studies may overestimate spore longevity in the environment. Spore settling rate was related to the nature of particulate material in the water column. Paraspores (or perhaps spores and toxin) of B. thuringiensis serovar israelensis (B.t.i.) had a greater tendency to adhere to and settle with suspended sediment and fine particulates than did paraspores of B. sphaericus. These observations may at least partially explain the greater persistence of B. sphaericus larvicidal activity in field tests than that of B.t.i..     

Host range and selected factors influencing the mosquito larvicidal activity of the PG-14 isolate of Bacillus thuringiensis var. morrisoni

Laboratory bioassay of the PG-14 isolate of Bacillus thuringiensis var. morrisoni (serotype 8a:8b) against early fourth instar larvae of 8 species of mosquitoes revealed a range of susceptibilities similar to the susceptibilities of these species to Bacillus thuringiensis var. israelensis (serotype 14). The most susceptible species were: Culex quinquefasciatus, Cx. salinarius, Anopheles albimanus and Aedes aegypti. The least susceptible species tested was An. quadrimaculatus. Separate bioassays of PG-14 against the four instars of Ae. aegypti demonstrated a strong negative correlation (R = -0.97) between larval age and susceptibility. Temperature significantly affected the stability of larvicidal toxin in aqueous suspensions of PG-14. Larvicidal activity of a bacterial suspension was nearly completely eliminated after 132 days of storage at 31 degrees C, but was essentially unchanged for those suspensions stored at 4 degrees C.     

Trial of Bacillus thuringiensis israelensis, a mosquito larvicide, against Porcellionides pruinosus, a terrestrial isopod

Woodlice, Porcellionides pruinosus, were not killed when exposed to granular Bacillus thuringiensis israelensis. Mortality was observed when woodlice received a topical application of the highest concentration of liquid Bti, but not at lower concentrations, nor when exposed via contaminated food or contaminated substrates.     

Field efficacy and nontarget effects of the mosquito larvicides temephos, methoprene, and Bacillus thuringiensis var. israelensis in Florida mangrove swamps

We compared the efficacy and nontarget effects of temephos, Bacillus thuringiensis var. israelensis (B.t.i.), and methoprene applied by helicopter to control mosquito larvae in mangrove swamps on Sanibel Island, FL, in May 1997. Three sites per treatment and 3 untreated sites were used. Temephos (Abate) was applied at 37 ml/ha (43% active ingredient [AI]), B.t.i. granules (Vectobac G) were applied at 5.606 kg/ha (200 International Toxic Units/mg), and methoprene (Altosid ALL) was applied at 213 ml/ha (5% AI). Efficacy was quantified by monitoring the survival of caged and uncaged larval Aedes taeniorhynchus. We quantified mortality of sentinel nontarget amphipods (Talitridae) at all sites, monitored the effect of temephos on flying arthropods using light traps, and collected dead insects in tarps suspended under mangroves in areas treated with either temephos or methoprene. Each pesticide showed good overall efficacy but occasional failures occurred. No detectable mortality of amphipods or flying insects attributable to pesticides was found. The inconsistent field efficacies of the pesticides indicate a need for reinspection of treated sites in this habitat.     

Factors influencing the activity of Bacillus thuringiensis var. israelensis treatments.

Environmental factors influence the effectiveness of microbial control agents in mosquito control programs. Four of these factors (water temperature, larval density, sunlight and the effect of associated filter feeders) were studied with Bacillus thuringiensis var. israelensis under laboratory and semifield conditions in Europe using different instars of Aedes vexans, Ae. aegypti and Culex pipiens. Bioassays conducted at a low temperature (5 degrees C) yielded 10-fold higher LC50 and LC90 values compared with those conducted at a high temperature (25 degrees C). The efficacy of B.t.i. decreased in a linear manner with increasing larval density. Sunlight can reduce the effectiveness of B.t.i. by several times. Competition in food intake by filter feeding Daphnia resulted in lower mortality of mosquito larvae after B.t.i. applications.     

Antibacteriophage action on the larvicidal activity of Bacillus thuringiensis H-14 and Bacillus sphaericus against Culex pipiens

An organically polluted mosquito breeding water was tested for the presence of bacteriophages which could inhibit the larvicidal activity of B. thuringiensis H-14 and B. sphaericus. More than one bacteriophage were isolated which could inhibit the tested bacteria. The sensitivity of the two bacterial species to 12 antibiotics was tested. Two of them, showed no antibacterial action, were selected and considered as antivirus agents in the bacteriophage assays; namely, Amoxycillin and Co-Trimoxazole. Results indicated their antivirus activity as the addition of the antibiotics to the isolated phages could permit normal bacterial growth as well as persistence of larvicidal activity against Culex pipiens larvae.     

An evaluation of Gambusia affinis and Bacillus thuringiensis var. israelensis as mosquito control agents in California wild rice fields

The mosquito control potential of the mosquitofish and Bacillus thuringiensis var. israelensis (Bti) were evaluated in experimental wild rice fields in Lake County, California. Fields were assigned one of six treatment: control, 1.1 kg/ha G. affinis, 3.4 kg/ha G. affinis, Bti only (6 kg/ha Vectobac granules), 1.1 kg/ha G. affinis plus Bti and 3.4 kg/ha G. affinis plus Bti. Gambusia affinis, at both release rates, significantly reduced the mosquito population at densities exceeding 100 fish/minnow trap. Treatments with Bti significantly reduced larval population; however, the populations in the fields without fish rebounded to pretreatment levels within two weeks. In fields stocked with G. affinis and treated with Bti, populations remained low after Bti treatment. Nontarget populations of arthropods were significantly lower in fields stocked with G. affinis than in fields without fish on one or more sampling dates.     

Field evaluation of new water-dispersible granular formulations of Bacillus thuringiensis ssp. israelensis and Bacillus sphaericus against Culex mosquitoes in microcosms.

A variety of formulations of Bacillus thuringiensis var. israelensis de Barjac (B.t.i.) and Bacillus sphaericus Neide (B.s.) have been studied for mosquito control under laboratory and field conditions. High efficacy, specificity, low risk of development of resistance, long shelf-life, and transportability, as well as the safety to nontarget organisms of these 2 microbial agents have been well documented. Some of the currently available formulations of B.t.i. and B.s. have low potency per unit mass. Research and development efforts are focusing on commercializing formulations with high potency and low minimum effective dosage that are suitable for long-distance shipment. To achieve this goal, new water-dispersible granule (WDG) formulations of both microbial agents were prepared and made available by Abbott Laboratories for evaluation. The newly developed WDGs of B.t.i. and B.s. with high potency dispersed readily in water with gentle agitation. These WDGs were evaluated and the minimum effective dosages were determined in microcosms against natural populations of Culex mosquitoes. The minimum effective dosage for B.t.i. WDGs with 4,000 International Toxic Units (ITU)/mg was 0.27-0.53 lb/acre which yielded significant control for up to 7-12 days. The minimum effective dosage for B.s. WDGs with 350-630 ITU/mg was 0.05-0.10 lb/acre, which yielded significant control of immature mosquitoes for up to 14-20 days.     

Bacterial control of mosquito larvae: investigation of stability of Bacillus thuringiensis var. israelensis and Bacillus sphaericus standard powders

Bacillus thuringiensis var. israelensis and Bacillus sphaericus products were assayed against their respective reference powders IPS82 and SPH88. Since their production in 1982 and 1988, the potency and larvicidal activity of these standard powders have been regularly checked on their test insects Aedes aegypti (for IPS82) or Culex pipiens (for SPH88). Over the 16-year evaluation period of IPS82 and 10-year evaluation period of SPH88, their potencies were considered stable. The global mean of each year's mean showed a coefficient of variation of less than 20%. Larval rearing was the most important factor in the reproducibility of the bioassay, although some variation also originated from the person performing the bioassay. This study demonstrated that the SPH88 standard could be kept in a stock suspension at 4 degrees C for 3 years without loss of potency. Moreover, after 9 years of storage in suspension, only a 2-fold decrease in the potency of SPH88 was detected.     

Laboratory study of the influence of water temperature and pH on Bacillus thuringiensis var. israelensis efficacy against black fly larvae (Diptera: Simuliidae)

An experimental formulation of Bacillus thuringiensis var. israelensis was used in the laboratory to assess the influence of water temperature and pH on the relationship between concentration, duration of exposure, and mortality of the northern black fly species Simulium decorum and Prosimulium mixtum/fuscum group. Mortality increases in both species with increases in duration of exposure, concentration, temperature and pH. Onset of death is shortened by increase in concentration and temperature. As temperature rises, the concentration of B.t.i. required to induce mortality decreases; the sharpest decline occurring between 12 and 18 degrees C for S. decorum, and between 4 and 8 degrees C for P. mixtum/fuscum larvae. Lower pH induces a loss of efficacy of the B.t.i. formulation on S. decorum larvae at 4 and 12 degrees C. Dialysis of the B.t.i. formulation at pH 11 for 2 h increases its potency against S. decorum larvae, suggesting an effect of an extralarval alkaline hydrolysis on the B.t.i. efficacy. An alkaline prehydrolysis of the paracrystalline bodies could therefore be used in cold and acidic environments to compensate for loss of efficacy.     

Differential efficacy evaluation of formulations of Bacillus thuringiensis Var israelensis against mosquitoes: a laboratory investigation

The efficacy evaluation of three formulations; wettable powder (W.P.) floating pellet and beads of Bacillus thuringiensis Var israelensis (Bti), revealed a greater susceptibility of the early larval instars of mosquitoes to Bti, sensitivity of anophelines to floating pellet, culicines to bead and equal efficacy and faster kill of W.P. to all the mosquitoes tested. A greater persistence of the slow release formulations, floating pellet and beads for 49 and 28 days against anophelines and culicines respectively was observed in contrast to a maximum persistence for 21 days in case of W.P. formulation.     

Floating bait formulations increase effectiveness of Bacillus thuringiensis var. israelensis against Anopheles larvae

The development and screening of floating-type bait formulations designed to improve the activity of bacterial toxins against larval Anopheles is described. Floating and spreading abilities of carrier particles (wheat flour) were compared using corn oil, lecithin, and two products yielding surface films on water (Arosurf and Liparol). Mixtures containing 1 or 5% Arosurf showed the best spreading abilities on a water surface, but strongly inhibited the ingestion of wheat flour by Anopheles albimanus larvae. Corn oil and lecithin improved spreading satisfactorily at a concentration of 5% and inhibited larval feeding by only 6-25%. To select a suitable concentration of active ingredient in formulations, Bacillus thuringiensis var. israelensis (B.t.i.) primary powder in concentrations ranging from 0.01 to 0.5% was mixed with wheat flour/corn oil mixtures and tested in quantities exceeding the gut volumes of treated larvae. Complete mortality was obtained with concentrations of 0.1% (Anopheles stephensi), 0.2% (An. albimanus), or 0.3% (An. quadrimaculatus) B.t.i. When in 175-liter containers the activity of formulations (5% corn oil, 0.2% B.t.i.) and of toxin suspensions was compared by conventional dosage-mortality regression, formulations were more active by a factor of 68 against An. stephensi, 39 against An. albimanus and 67 against An. quadrimaculatus.     

Mosquito larvicidal activity of active constituent derived from Chamaecyparis obtusa leaves against 3 mosquito species

Mosqutio larvicidal activity of Chamaecyparis obtusa leaf-derived materials against the 4th-stage larvae of Aedes aegypti (L.), Ochlerotatus togoi (Theobald), and Culex pipiens pallens (Coquillett) was examined in the laboratory. A crude methanol extract of C. obtusa leaves was found to be active (percent mortality rough) against the 3 species larvae; the hexane fraction of the methanol extract showed a strong larvicidal activity (100% mortality) at 100 ppm. The bioactive component in the C. obtusa leaf extract was characterized as beta-thujaplicin by spectroscopic analyses. The LC50 value of beta-thujaplicin was 2.91, 2.60, and 1.33 ppm against Ae. aegypti, Oc. togoi, and Cx. pipiens pallens larvae. This naturally occurring C. obtusa leaves-derived compound merits further study as a potential mosquito larval control agent or lead compound.     

Bti微胶囊制剂防治蚊虫的研究

苏云金杆菌以色列变种（Ｂｔｉ）广泛应用于现场控制蚊幼虫，取得良好的杀灭效果。但由于现有剂型持效期短，限制了更大规模的应用，也给储存运输带来麻烦。本研究主要在室内评价了不同浓度Ｂｔｉ微胶囊制剂对中华按蚊、淡色库蚊和白纹伊蚊幼虫的效果及持效。结果表明，该种制剂有一定的漂浮性，在水内稳定性较好，具较长的持效期，１．２５ｐｐｍ浓度４周内对白纹伊蚊幼虫仍有较理想的杀灭效果。推荐防治中华按蚊、淡色库蚊和白纹伊蚊的需要剂量分别为０．２５、１．２５和０．０５ｐｐｍ，每２～３周施药１次。     

Efficacy of two formulations of Bacillus thuringiensis var. israelensis (H-14) against Aedes vexans and safety to non-target macroinvertebrates

An experimental Sandoz formulation of Bacillus thuringiensis var. israelensis (SAN 402 SC 98) was several times more effective than Abbott ABG 6188 against larvae of Aedes aegypti and Ae. vexans in the laboratory. Field applications of SAN 402 SC 98 at 0.25 liter/ha and ABG 6188 at 1.00 liter/ha resulted in more than 97% control of Ae. vexans larvae after 48 hours, with residual activity of 24 hours or less. The amphipod, Hyallela azteca, and 4 species of water beetles were apparently unaffected by 100 ppm SAN 402 SC 98, which is 1,449 times the 24 hour LC50 for third instar Ae. vexans larvae.     

Effects of algae on the efficacy of Bacillus thuringiensis var. israelensis against larval black flies

Personnel from several black fly control programs have reported that the efficacy of Bacillus thuringiensis var. israelesis (Bti) is reduced during periods when algal concentrations are high in the waterways. Although the reduction in Bti-induced mortality in black fly larvae is presumed to be related to the presence of algae, no scientific data support this theory. In this study, 4 genera of algae (Microcytis, Scenedesmus, Dictrosphaerium, and Chlorella) commonly detected in Pennsylvania rivers where Bti-induced mortality in black fly larvae has been reduced were assessed to determine their respective effects on Bti-induced mortality by using an orbital shaker bioassay with laboratory-reared black fly larvae (Simulium vittatum cytospecies IS-7). A significant reduction in Bti-induced mortality was observed when Scenedesmus was present in the flasks at concentrations > or = 16,000 cells/ml. The Bti-induced mortality of larvae was not significantly reduced when Chlorella, Dictyosphaerium, or Microcytis was present in the flasks, even at concentrations > or = 250,000 cells/ml. These results indicate that the presence of certain types of algae can reduce the mortality of black flies exposed to Bti. Although not clearly defined, the mechanisms involved may be related to algal morphology due to overall size and structures associated with certain types of algae, and possible interference with feeding.