Factors Affecting Residual Dosages of Two Formulations of Bacillus thuringiensis subsp. israelensis Tested in the Same Stream During a 3-Year Experiment

Although many field trials have been conducted using Bacillus thuringiensis subsp. israelensis (Bti)-based formulations, most have been in rivers with different biotic and abiotic conditions thus rendering the evaluation of their performance very difficult. Recently, results of a threeyear experiment using a new field procedure brought new insight into the behavior and the performance (carry) of two liquid formulations of Bti, Teknar HP-D and Vectobac 1200L, tested in the same lotic environment and under similar abiotic and biotic conditions. Factors such as discharge, water temperature and the hyporheic zone were identified as elements affecting the downstream loss of activity of both products. However, to better understand the reduction of black fly mortality along a stream (measured by using gutters), data of residual dosages of both products (measured by laboratory assay with mosquito larvae) were compared with reduction of black fly mortality. Bti toxic activity was monitored from water samples taken at different distances downstream from an application point, and from probes driven into the hyporheic zone, to study the effects of abiotic factors on the loss of the toxic crystals. Results showed that the loss of dosage was exponential for both products but more crystals were recovered from Vectobac 1200L along the stream than from Teknar HP-D. However, the latter was more efficient, i.e. less toxins were needed to kill 50% of black fly larvae both in temperate (16°C) and warmer (19.5-22°C) water. Also, a rise in water temperature had a greater effect in the kill induced by Vectobac 1200L compared to Teknar HP-D. For the same residual dosages present at the stations, longer carries of toxin activity (higher mortalities) were obtained in warmer water. Finally, the hyporheic zone was identified as a major source of loss of activity of Bti products. Large stream discharges decreased the effect of the hyporheic zone and that was reflected in longer carry of the products.     

Influence of Stream Profile and Abiotic Factors on the Performance and Residual Dosages of Two Commercial Formulations of Bacillus thuringiensis subsp. israelensis During a 2-Year Experiment

Recently, results of a 3-year study showed the importance of abiotic factors such as water temperature, stream discharge and the hyporheic zone on the behaviour, performance and loss of the residual dosages (amount of the injected dosage left at each station) of two commercial liquid formulations of Bacillus thuringiensis subsp. israelensis ( Bti ), Teknar HP-D and Vectobac 1200L. Experiments were performed in the same stream and under environmental conditions that allowed comparisons of results between formulations. To improve our understanding of the influence of abiotic factors on the behaviour and the performance of these formulations, a 2-year experiment using the same products was conducted in another river characterized by a much different profile, higher discharges and colder temperatures. A field procedure based on a system of gutters located on the bank of the river already used in the previous 3-year experiment was utilized. Black fly larval mortalities were recorded along the river at different distances (stations) to evaluate the behaviour and the performance of the products while water samples taken at the stations were tested in the laboratory against mosquito neonate larvae to evaluate the residual dosages of Bti at each station. Results showed that higher residual dosages of the Vectobac 1200L were recovered compared to Teknar HP-D along the river. Although higher dosages were recovered, higher mortality was observed only for the Vectobac 1200 L in high discharge conditions. As seen before, the hyporheic zone (interstitial water between the streambed and groundwater) produced a major loss of the dosages in the first meters of the river. But because of the river profile, the hyporheic zone had a lesser effect on the loss of the dosages further in the river resulting in very long carries for both Teknar HP-D and Vectobac 1200L.     

Storage Stability of Two Liquid Formulations of Bacillus thuringiensis subsp. israelensis and Effect of Freezing Over Time

Over the last two decades, many tests have been performed in the field to investigate the behaviour (persistence, carry, loss of activity, etc.) of different Bacillus thuringiensis subsp. israelensis ( Bti ) formulations. Depending on the experimental protocols, a single container of a formulation could be used more than once over time and field samples containing Bti may have to be frozen to preserve them for bioassays to be performed later. Thus, it is necessary to know how long a formulation could keep its level of efficacy and also the effects of time on frozen samples. Our results showed that the efficacy of two commercial liquid formulations of Bti (Teknar HP-D and Vectobac 1200L) when tested against Aedes triseriatus behaved differently over time when kept at room temperature. Teknar HP-D remained stable for the first two years and its LC 50 increased by 20% the third year. For Vectobac 1200L, although its larvicidal activity was better than that of Teknar HP-D every year, there was an increase in LC 50 by 22% the second year and by another 20% the third year for a total loss of activity of 46% over the three-year study. The efficacy of suspensions made with both formulations was greatly affected by freezing and the loss of efficacy increased over time. About half of the efficacy of Teknar HP-D was lost after one week of freezing and stayed at that level for three months, while with Vectobac 1200L, no significant effect of freezing was seen after one month, when compared to fresh material. However, both products showed similar efficacy after three or six months of freezing. Overall, the LC 50 s of both products had increased by a factor of about 2.5 after six months of freezing.     

A New Field Procedure and Method of Analysis to Evaluate the Performance of Bacillus thuringiensis subsp. israelensis Liquid Formulations in Streams and Rivers

Many field tests have shown Bacillus thuringiensis subsp. israelensis ( Bti ) to be an effective simuliid larvicide. However, literature indicates that an effective evaluation and comparison of Bti -based formulations when tested in streams or rivers is difficult. Most field trials have been conducted in different rivers (different discharge, river profile, water temperature, suspended matter, larval species, etc.), thus rendering the evaluation of the performance of liquid formulations of Bti very arbitrary or even impossible. A new field procedure is proposed to evaluate the performance of liquid formulations of Bti in the same lotic environment and under similar abiotic and biotic conditions. The procedure, based on a system of gutters located on the bank of a stream, showed very good reproducibility of the mortality levels of the target pest(s) recorded at various distances (stations) along the stream over a two-year field trial, proving the efficacy of the system. The system allows either a single formulation or different formulations to be tested repeatedly in the same portion of a stream, thereby providing a more accurate evaluation of the performance of a Bti formulation or a much better comparison between different formulations. The use of the probit model (allowing comparison of slopes and intercepts) gives a reliable statistical value for the analysis of the results. Moreover, this system is not expensive and can be transferred easily to other streams or rivers.     

Factors affecting the efficacy of three commercial formulations of Bacillus thuringiensis var. israelensis against species of European black flies

The efficacy of three commercial formulations of Bacillus thuringiensis var. israelensis against two European black fly species was determined. The effect of seston concentration, temperature and larval size on the efficacy of the formulations was also determined. Teknar‐WDC was found to be almost four times as effective as Bactimos‐WP though only slightly more efficient than Vectobac‐AS against Simulium variegatum gp. larvae. Small larvae ( < 2.5 mm) were more susceptible to both Teknar‐WDC and Vectobac‐AS than large larvae ( > 5 mm) though no difference was seen with Bactimos‐WP. High seston concentrations both before and during bioassays significantly decreased the efficacy of all formulations. Increased temperatures increased the efficacy of all formulations.     

Laboratory and field evaluations of industrial formulations of Bacillus thuringiensis serovar. israelensis against some mosquito species of central Italy

Two wettable powder formulations (Bactimos and Vectobac) and a flowable concentrate (Teknar) formulation of Bacillus thuringiensis serovar. israelensis were evaluated as larvicides of Culex pipiens, Aedes caspius, and Aedes detritus. In the laboratory, the levels of susceptibility of these species to the test formulations were essentially similar and corresponded to their relative potencies; the LC₉₀ values ranged from 0.042 to 0.37 ppm. C. pipiens, A. caspius, and A. detritus, in that order, were susceptible to the biocide. Under field conditions in central Italy. Bactimos at 0.5 kg/ha gave 98% control of C. pipiens in an irrigation canal. Teknar at 1.25 liters/ha gave 86–100%, and at 2.5 liters/ha gave 90–100% control of C. pipiens in two natural ponds. Against A. caspius in salt marsh habitats, Bactimos at 0.5 kg/ha and Teknar at 2.5 liters/ha yielded complete control of the larvae, while a lower rate (0.2 kg/ha) of Bactimos, and Vectobac at 0.5 kg/ha resulted in 82–94% and 67–91% control, respectively. Higher rates (0.75 and 1.0 kg/ha) of Vectobac gave 76–100% and 98–100% control of A. caspius. Bactimos at 0.15 kg/ha gave 93–98% control of A. detritus in two salt marsh ponds. B. thuringiensis serovar. israelensis is practically economical for the control of C. pipiens, A. caspius, and A. detritus in the various biotopes in central Italy.     

Low Persistence of Bacillus thuringiensis Serovar israelensis Spores in Four Mosquito Biotopes of a Salt Marsh in Southern France

We studied the persistence of Bacillus thuringiensis serovar israelensis (Bti) in a typical breeding site of the mosquito Ochlerotatus caspius in a particularly sensitive salt marsh ecosystem following two Bti-based larvicidal applications (Vectobac 12AS, 1.95 L/ha). The treated area was composed of four larval biotopes that differed in terms of the most representative plant species (Sarcocornia fruticosa, Bolboschoenus maritimus, Phragmites australis, and Juncus maritimus) and the physical and chemical characteristics of the soil. We sampled water, soil, and plants at various times before and after the applications (from spring to autumn, 2001) and quantified the spores of B. thuringiensis (Bt) and Bacillus species. The B. cereus group accounted for between 0% and 20% of all Bacillus spp. before application depending on the larval biotope. No Bti were found before application. The variation in the quantity of bacilli during the mosquito breeding season depended more on the larval biotope than on the season or the larvicidal application. More bacilli were found in soil (10(4)-10(6) spores/g) than on plant samples (10(2)-10(4) spores/g). The abundance in water (10(5) to 10(7) spores/L) appeared to be correlated to the water level of the breeding site. The number of Bti spores increased just after application, after declining; no spores were detected in soil or water 3 months after application. However, low numbers of Bti spores were present on foliage from three of the four studied plant strata. In conclusion, the larvicidal application has very little impact on Bacillus spp. flora after one breeding season (two applications).     

The effect of seston on mortality of Simulium vittatum (Diptera: Simuliidae) from insecticidal proteins produced by Bacillus thuringiensis subsp. israelensis

Water was collected from a site on the Susquehanna River in eastern Pennsylvania, where less-than-optimal black fly larval mortality had been occasionally observed after treatment with Bacillus thuringiensis subsp. israelensis de Barjac insecticidal crystalline proteins (Bti ICPs). A series of experiments was conducted with Simulium vittatum Zetterstedt larvae to determine the water related factors responsible for the impaired response to Bti ICPs (Vectobac 12S, strain AM 65-52). Seston in the water impaired the effectiveness of the ICPs, whereas the dissolved substances had no impact on larval mortality. Individual components of the seston then were exposed to the larvae followed by exposure to Bti ICPs. Exposure of larvae to selected minerals and nutritive organic material before ICP exposure resulted in no significant decrease in mortality. Exposure of larvae to silicon dioxide, cellulose, viable diatoms, and purified diatom frustules before Bti ICP exposure resulted in significant reductions in mortality. Exposure of larvae to purified diatom frustules from Cyclotella meneghiniana Kützing resulted in the most severe impairment of mortality after Bti ICP exposure. It is postulated that frustule-induced impairment of feeding behavior is responsible for the impairment of larval mortality.     

Influence of molting on efficacy of two functionally different larvicides: Bti and temephos

The sensitivity of the second, third (L3), and fourth (L4) instars of Culex pipiens L. and Ochlerotatus sticticus (Meigen) to two larvicides, temephos and Bacillus thuringiensis variety israelensis (Bti) was tested with and without molting occurring in the experiments. In the first experiment for both tested mosquito species, LC50 values increased from the second to the fourth instar when tested against temephos, whereas for Bti the opposite trend was observed. The highest LC50 value was found for fourth instars of Cx. pipens (0.00405 mg/liter) against temephos and for second instars of Oc. sticticus (0.267 mg/liter) against Bti. The determined LC50 values for the second and third instars of both species decreased with an increased number of molted larvae in the experiments with temephos. For the experiments with Bti molting did not have any significant influence on LC50 values, except a small increase in toxicity during the L3/L4 molt of Oc. sticticus. These findings could help assess and define larviciding, as well as influence the quantity of larvicides needed for an efficient treatment.     

Effects of Bacillus thuringiensis var. israelensis on Target and Nontarget Organisms: a Review of Laboratory and Field Experiments

Since the discovery of Bacillus thuringiensis var. israelensis (Bti) in 1976, extensive literature has proved its efficacy to control mosquitoes and black flies, of which many species are known as important vectors of diseases or simply as pests of humans and animals. Since 1978, Bti has been used in many countries on all continents and numerous studies have been made on target mosquitoes and black flies, as well as nontarget organisms (NTO). This review analyses the results of 75 studies on these organisms covering approximately 125 families, 300 genera and 400 species. Different factors such as species, instar, feeding behaviour and environmental parameters (larval density, water temperature, suspended matter etc.) may drastically affect the efficacy of the Bti products. This is addressed in detail by reviewing the main factors affecting mosquitoes as well as black flies. The results of a wide range of laboratory and field experiments using different target and nontarget species, various preparations and formulations of Bti and different biotic or abiotic factors are present in the literature, making the data difficult to compare on a common basis. Our analysis shows that, under different application conditions, the effects of Bti on target and nontarget organisms may be hard to predict. Although Bti has been proclaimed to be relatively highly specific, some studies show that some NTO are affected either by single or repeated Bti treatments. Present use against black flies seems ecologically acceptable. High frequencies of application and/or overdosages against mosquitoes may result in some persistence of the toxin crystals and ultimately this may have adverse effects on the food web. A long-term study (published in 1998) in mosquito habitats has shown that intensive Bti treatments over three years did in fact produce an impact on the food web in wetlands. This raises questions, for the first time, on Bti environmental specificity. The importance of this impact is discussed and the alternatives for practical pest control are considered. Some modifications of Bti use against mosquitoes, guided by research, is probably the best of these alternatives.     

In vitro assay for testing the toxicity of Bacillus thuringiensis subsp. israelensis delta endotoxin using blood agarose plates

An in vitro assay system that used erythrocytes was developed to assess the toxicity of delta endotoxin of Bacillus thuringiensis subsp. israelensis (Bti), a mosquito larvicidal agent. This endotoxin was activated at high alkaline pH without causing alkaline injury to the erythrocytes. The assay is carried out on blood agarose plates, prepared in MOPS-buffered saline (0·01 mol 1^-1), pH 7·0. The threshold dose for toxicity of Bti HD-567 was 8 ng protein and that of Bti H-14 and IPS-82 standard were 6 ng and 4 ng protein respectively. The assay is rapid, sensitive, quantitative and applicable to different Bti formulations to assess the toxicity of the products. Only the activated toxin is detected by this method and has good correlation with insect bioassay. The percentage toxicity dose of different Bti preparations agrees well with percentages of LC₅₀ values when the second instar larvae of Aedes aegypti were tested.     

The bacterium, Lysinibacillus sphaericus, as an insect pathogen

Since the first bacteria with insecticidal activity against mosquito larvae were reported in the 1960s, many have been described, with the most potent being isolates of Bacillus thuringiensis or Lysinibacillus sphaericus (formerly and best known as Bacillus sphaericus). Given environmental concerns over the use of broad spectrum synthetic chemical insecticides and the evolution of resistance to these, industry placed emphasis on the development of bacteria as alternative control agents. To date, numerous commercial formulations of B. thuringiensis subsp. israelensis (Bti) are available in many countries for control of nuisance and vector mosquitoes. Within the past few years, commercial formulations of L. sphaericus (Ls) have become available. Because Bti has been in use for more than 30 years, its properties are well know, more so than those of Ls. Thus, the purpose of this review is to summarise the most critical aspects of Ls and the various proteins that account for its insecticidal properties, especially the mosquitocidal activity of the most common isolates studied. Data are reviewed for the binary toxin, which accounts for the activity of sporulated cells, as well as for other toxins produced during vegetative growth, including sphaericolysin (active against cockroaches and caterpillars) and the different mosquitocidal Mtx and Cry toxins. Future studies of these could well lead to novel potent and environmentally compatible insecticidal products for controlling a range of insect pests and vectors of disease.     

Field evaluation of four biorational larvicide formulations against Anopheles albimanus in Honduras

Four biorational larvicide formulations: Teknar (B.t.i.), Arosurf MSF (Monomolecular Surface Film), Arosurf MSF combined with Teknar, and SAN-809-I ([s]-methoprene combined with B.t.i.), were evaluated against naturally occurring populations of the malaria vector mosquito Anopheles albimanus Wiedemann in Honduras. All formulations reduced the mean number of larvae per sample area to 0 within 48 h after treatment, and gave significant (P less than 0.05) control when compared with similar untreated areas for at least 10 days after treatment. It is concluded that each of these four formulations can be used effectively to control the larvae of An. albimanus in Honduras.     

Laboratory and field evaluation of formulated Bacillus thuringiensis var. israelensis as a feed additive and using topical applications for control of Musca domestica (Diptera: Muscidae) larvae in caged-poultry manure

Infestations of house flies, Musca domestica L., are a continual problem around poultry establishments. Acute toxicity of two commercial Bacillus thuringiensis variety israelensis (Bti) formulations (water-dispersible granules and bran formulation) was evaluated against larvae in the laboratory and against natural populations of M. domestica larvae in the field applied in feed to chickens and as topical applications in the poultry houses. Bioassay data showed that susceptibility of M. domestica larvae increased to a given concentration of Bti as the duration of exposure increased. In the laboratory studies, the LC(50) values of Bti for the larvae ranged between 65 and 77.4 μg/ml. In the field, a concentration of 10 g Bti/kg of feed resulted in 90% reduction of larvae at 4 wk after treatment. A higher concentration (2 g/liter) of Bti in spray applications was not significantly more effective than the lower concentration of 1 g/liter. Adding Bti to chicken feed is potentially an efficient measure for the management and control of house flies in caged-poultry facilities.     

The naturally derived insecticide spinosad is highly toxic to Aedes and Anopheles mosquito larvae

Spinosad is a naturally derived biorational insecticide with an environmentally favourable toxicity profile, so we investigated its potency against mosquito larvae (Diptera: Culicidae). By laboratory bioassays of a suspension concentrate formulation of spinosad (Tracer), the 24 h lethal concentration (LC50) against Aedes aegypti (L.) third and fourth instars was estimated at 0.025 p.p.m. following logit regression. The concentration-mortality response of third- and fourth-instar Anopheles albimanus Weidemann did not conform to a logit model. The LC50 value of spinosad in Anopheles albimanus was 0.024 p.p.m. by quadratic linear regression. A field trial in southern Mexico demonstrated that spinosad 1 p.p.m. compared with the standard temephos (Abate) 1% granules 100 g/m3 water prevented Ae. aegypti breeding in plastic containers of water for 8 weeks; at 10 p.p.m. spinosad prevented breeding for > 22 weeks. In another field trial, spinosad at 5 p.p.m. and temephos both completely eliminated reproduction of Ae. aegypti for 13 weeks. In contrast, the bacterial insecticide Bacillus thuringiensis var. israelensis (Bti, Vectobac) AS) performed poorly with just 2 weeks of complete inhibition of Ae. aegypti breeding. Spinosad also effectively prevented breeding of Culex mosquitoes and chironomids in both trials to a degree similar to that of temephos. We conclude that spinosad merits evaluation as a replacement for organophosphate or Bti treatment of domestic water tanks in Mesoamerica. We also predict that spinosad is likely to be an effective larvicide for treatment of mosquito breeding sites.     

Spray-dried Bacillus thuringiensis Serovar israelensis formulations for control of Aedes aegypti larvae

Suspensions containing 0.25 and 1.25 g/liter of Bacillus thuringiensis subsp. israelensis (Bti) spore-toxin complex were spray-dried by using maltodextrin DE-6, corn starch, and nixtamalized corn flour (25 g/liter) as materials to entrap active delta-endotoxin. The inlet air temperature of the drier was kept constant at 141 degrees C and the outlet temperature was maintained at 60 or 70 degrees C. The Probit analysis of the concentration-mortality response of third instars of Aedes aegypti (L.) larvae of the spray-dried products at 60 degrees C showed that LC50 values for maltodextrin DE-6 with 1 and 5% spore-toxin complex were 4 and 10% higher in toxicity, respectively, than that for the unformulated spore-toxin complex without drying. The LC50 value for corn starch with 1 and 5% of spore-toxin complex were also higher in toxicity (7 and 8% respectively). However, LC50 values for nixtamalized corn flour with one and 5% spore-toxin complex were 81 and 55% higher in toxicity, respectively. Dried products contain an a(w) < or = 0.7, suggesting that they are able to keep the products without microorganism growth for longer periods. The scanning electron microscope of Bti spray-dried formulations with nixtamalized corn flour showed smooth spherical particles entrapping the active ingredient. These results suggested that Bti spore-toxin complex formulated with maltodextrin DE-6, corn flour, and nixtamalized corn flour, and then spray-dried may increase larval feeding and thus increase activity against Ae. aegypti larvae.     

Biologically based insecticides for the control of immature Australian mosquitoes: a review

Abstract  Our review outlines changes in insecticide usage for mosquito control against Australian mosquitoes, with a focus on biologically based insecticides. The most widely used microbial insecticide is Bacillus thuringiensis var. israelensis de Barjac ( Bti ) and it is regularly used in estuaries and freshwater. In comparison, the use of Bacillus sphaericus (Neide) is limited to freshwater. The only insect growth regulator registered for mosquito control in Australia is s -methoprene, and it is used in both estuaries and freshwater. Although biologically based insecticides have been used for more than 10 years, there remains knowledge gaps surrounding the use of specific formulations in specific habitats: for example, VectoBac WG ( Bti ) and VectoBac G ( Bti ) in containers containing freshwater, or Altosid pellets ( s -methoprene) in ground-pool freshwater habitats. Where broad-scale mosquito control programs have been implemented in Australia, a reduction in the incidence of mosquito-borne disease, mainly Ross River virus, has been recorded. Therefore, the future application of these products is supported. The use of insecticides should be integrated with public education, biological control, physical habitat modification and coupled with spatial risk assessment.     

Inversion polymorphism in malaria mosquito Anopheles messeae. Part X. Resistance of larvae with different genotypes to toxins of crystal-forming bacteria Bacillus thuringiensis subsp. israelensis (serovar H14)

Insensibility of larvae with different chromosomal inversions to the toxins of Bacillus thuringiensis subsp. israelensis (Bti) was examined. It has been shown that larvae with inversion combinations XL0(XL1)2R0-3R0-3L0 had greater variability after treatment with Bti than larvae with inversions XL1(XL2)2R1-3R1-3L1(3L0). The former inversion combinations were previously shown to dominate in the south of the species area and to be supported by K-selection. The latter inversion combinations form "northest" karyotypes and are supported by r-selection. It is obvious that genetic effects of treatments with Bti depend on the population's structure and the directions of natural selection. The decrease in the level of heterozygotes after treatment of larvae with Bti reflects destruction of the system of genetic homeostasis in the natural populations of A. messeae.     

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.     

Quality control of Bacillus thuringiensis ssp. israelensis products based on toxin quantification with monoclonal antibodies

Quality control of Bacillus thuringiensis ssp. israelensis (Bti) products is currently based on international toxic units (ITUs). The potency of products is related to the activity of a standard (IPS-82, Institute Pasteur, Paris) assessed in bioassays using Aedes aegypti as a target host. The procedure is time consuming and costly, often producing variable results. The activity of Bti is based on four different insecticidal crystal proteins (ICPs): Cry4Aa, Cry4Ba, Cry11Aa and Cyt1Aa. Monoclonal antibodies were produced using IPS-82 for immunisation and an enzyme-linked immunosorbent assay (ELISA) was developed. Antibodies were selected with specificity against Cry11A and Cyt1A. Cry4 specific antibodies could not distinguish between Cry4A and Cry4B. Within five replicate assessments of the three ICPs (in µg mg^-1 ICP protein), an error between 3 and 8% was recorded, whereas a 14% error was obtained comparing seven samples of the same production batch for ITUs mg^-1. The toxicity against A. aegypti expressed in ITUs correlated well with the results of the ELISA (correlation coefficient r Cyt 1=0.79; Cry 11=0.87; Cry 4=0.91) also when related to the sum of all ICPs (r=0.87). The ELISA can reduce efforts to determine Bti quality compared with the labour-intensive and variable ITU bioassay.