Coumaphos Residues Research Articles

Transcriptional responses of detoxification genes to coumaphos in a nontarget species, Galleria mellonella (greater wax moth) (Lepidoptera: Pyralidae), in the beehive environment

The greater wax moth Galleria mellonella is a cosmopolitan pest of hives of the western honey bee Apis mellifera, where it remains exposed to varroicides applied by beekeepers in past decades as pest management chemicals for control of Varroa destructor, a devastating ectoparasite of bees. The prolonged presence of coumaphos residues, an organophosphate varroicide, in beeswax may be responsible for current levels of tolerance exhibited by G. mellonella, a non-target species that infests beehives. In this study, a field-collected strain of waxworms exhibited a higher LC50 value for coumaphos than that of a laboratory strain that had not been continuously exposed to coumaphos residues at field concentrations. Despite its higher tolerance for coumaphos, the field strain experienced growth inhibition at ecologically relevant concentration of coumaphos. Moreover, at low environmental concentrations that did not alter growth, detoxification gene expression levels were substantially altered. RNA-Seq analysis revealed 1181 and 658 differentially expressed genes in fat body and midgut, respectively, with 378 and 186 of those genes upregulated. This large-scale upregulation encompassed 21 genes encoding cytochrome P450 monooxygenases (CYPs), 13 encoding UDP-glycosyltransferases (UGTs), 5 encoding glutathione-S-transferases (GSTs), 2 encoding carboxylesterases (COEs), and 2 encoding ABC transporters (ABCs) in either tissue. Expression analysis of 13 selected candidate detoxification genes by RT-qPCR was consistent with their expression from RNA-Seq data. In sum, our results indicate that long-lasting pesticide residues in beeswax from past Varroa mite management may continue to act as selective agents on detoxification systems of hive residents other than the initial target species and that multiple resistance mechanisms to these chemicals may coexist within the beehive fauna.

Toxicity of Coumaphos Residues in Beeswax Foundation to the Honey Bee Brood.

Coumaphos is one of the most frequently detected pesticides in recycled beeswax. The objective was to assess the maximal level of coumaphos in foundation sheets that could exist without lethal effects on the honey bee larvae. Brood development was followed in cells drawn on foundation squares containing coumaphos ranging from 0 to 132 mg/kg. Furthermore, larval exposure was determined by measuring the coumaphos level in the drawn cells. Coumaphos levels in the initial foundation sheets up to 62 mg/kg did not increase brood mortality because the emergence rates of bees raised on these foundation squares were similar to controls (median of 51%). After a single brood cycle, coumaphos levels in the drawn cells were up to three times lower than the initial levels in foundation sheets. Hence, coumaphos levels of 62 mg/kg in the initial foundation sheets, almost the highest exposures, resulted in levels of 21 mg/kg in drawn cells. A significantly reduced emergence rate (median of 14%) was observed for bees raised on foundation sheets with initial coumaphos levels of 132 mg/kg, indicating increased brood mortality. Such levels resulted in coumaphos concentrations of 51 mg/kg in drawn cells, which is close to the median lethal concentration (LC50) as determined in previous in vitro experiments. In conclusion, brood mortality was increased on wax foundation sheets with initial coumaphos levels of 132 mg/kg, while no elevated mortality was observed for levels up to 62 mg/kg. Environ Toxicol Chem 2023;42:1816-1822. © 2023 The Authors. Environmental Toxicology and Chemistry published by Wiley Periodicals LLC on behalf of SETAC.

First national survey of residues of active substances in honeybee apiaries across Spain between 2012 and 2016.

This nationwide monitoring aimed to investigate the prevalence of residues of plant protection products (PPPs) and veterinary medicine products (VMPs) based on random selection of apiaries of Apis mellifera. For a three-year period (2012, 2013 and 2016), this study targeted 306 PPPs, VMPs and other active substances in 442 samples of bee bread honeycomb (BBHC) and 89 samples of honeybees collected from up to 177 apiaries. The results indicate that honeybees were most often exposed to residues of coumaphos, tau-fluvalinate, chlorfenvinphos, and acrinathrin, with a prevalence from a maximum of 98.8% to 49.4% in BBHC samples. Residues of coumaphos, tau-fluvalinate, amitraz (DMF + DMPF), carbendazim and orthophenylphenol were also frequently detected, from a maximum of 55.1% to 13.5% of the honeybee samples. Neonicotinoid residues, namely clothianidin and thiamethoxam, whose outdoor uses in crops are completely banned in EU, were not detected. Imidacloprid was found in 3.4% to 13.3% of samples during 2013 and 2016, respectively. Imidacloprid exceeded its acute toxicity (LD50) value for honey bees in two samples of BBHC. Fipronil was detected in 0.5% of the samples during 2013. The diversity of active substances found (% of different residues analyzed) ranged from 33.9% to 37.2% in BBHC from 2012, 2013 to 2016, and was of 26.5% in honeybees in 2016. In at least 54% of the samples, the total residue load was in the range of 200 to 1500 μg·kg-1. Up to 50% of BBHC samples were positive for one or two residues. No toxic residues for honeybees were detected in up to 88.8% of bee samples. This systematic surveillance of active substances assisted the evaluation of which target pesticides to look for and provided support to the competent authorities in the bee health decision-making.

Investigation on amitraz, coumaphos and thymol concentrations in honey produced by Slovenian beekeepers in 2020

A survey on concentrations of veterinary drug residues amitraz, coumaphos and thymol in honey, produced in year 2020 by Slovenian beekeepers, was conducted. 100 samples were analysed: 22 from organic and 78 from conventional production, with two analytical methods. In method for determination of coumaphos and thymol samples were extracted with acetone, petroleumether and dichlorometane. In method for determination of amitraz and its degradation products, samples were hydrolisated with HCl and NaOH, extractied with n-hexane and derivatisated with heptafluorobutyric anhydride. Determination in both methods was performed with gas chromatograph coupled with mass spectrometer. Measured concentrations of amitraz, coumaphos and thymol were in the range of 0.01-0.12 mg kg-1, 0.02-0.06 mg kg-1 and 0.08-0.17 mg kg-1, respectively. In 61 % of samples analysed no residues of amitraz, thymol and coumaphos were found. Data obtained were compared with the data from literature. Chronic and acute exposure were calculated for consumers. Maximum chronic exposure for amitraz and thymol was 0.1 % and 0.05 % of acceptable daily intake, respectively. Maximum acute exposure for amitraz and thymol was 4 % and 0.8 % of acute reference dose, respectively

Coumaphos residues in beeswax after a single application of CheckMite® affect larval development in vitro

Coumaphos residues in beeswax after a single application of CheckMite® affect larval development in vitro

Chronic toxicity of amitraz, coumaphos and fluvalinate to Apis mellifera L. larvae reared in vitro

The effects of chronic exposure to common acaricides on Apis mellifera survival, developmental rate and larval weight were tested in the laboratory. Larvae were reared in vitro and fed a diet containing amitraz: 1.5, 11, 25 and 46 mg/L; coumaphos: 1.8, 6, 8 and 25 mg/L; or fluvalinate: 0.1, 1, 2.4 and 6 mg/L. The dependent variables were compared for groups feeding on treated diets and control diets: positive control, 45 mg/L dimethoate; solvent control; and negative control. Bee survival decreased in the 46 mg/L amitraz and 25 mg/L coumaphos treatments but not in any fluvalinate treatment. Furthermore, the developmental rate decreased in individuals treated with 46 mg/L amitraz. In our study, larvae exposed to acaricides at concentrations similar to maximum residue in pollen and honey/nectar had no detectable change in survival or developmental rate. Given that pollen and honey/nectar represent only a small part of larval diet, we suggest that residues of amitraz, coumaphos and fluvalinate at the levels we tested are unlikely to impact immature worker bee survival in the field, though our data do not preclude any sublethal effects that may result from bee exposure to these compounds or possible synergisms when they co-occur in bee colonies.

Coumaphos residues in honey, bee brood, and beeswax after Varroa treatment

Residues of acaricide coumaphos were assessed in honey, bee brood, and beeswax during a 2-year field experiment. Honey, bee brood, and beeswax samples were collected before and after routine use of coumaphos in the treatment of bee colonies against varroosis in two consecutive years. Determination of coumaphos in honey and bee brood was based on RP-HPLC with UV detection after a liquid-liquid extraction with hexane or ethyl acetate. Coumaphos in beeswax was identified and quantified by GC/MS. Results indicate the undetectable presence of coumaphos in honey. In bee brood, coumaphos was observed after the treatment. In beeswax, the accumulation of coumaphos was determined not only in hives where it was used but also in hives nearby in which coumaphos was not used. Results indicate the accumulation of coumaphos in bee brood and beeswax. Due to the coumaphos accumulation this drug should be used only in strongly affected bee colonies.

Boron and Coumaphos Residues in Hive Materials Following Treatments for the Control of Aethina tumida Murray.

In the search of alternatives for controlling Aethina tumida Murray, we recently proposed the BAA trap which uses boric acid and an attractant which mimics the process of fermentation caused by Kodamaea ohmeri in the hive. This yeast is excreted in the feces of A. tumida causing the fermentation of pollen and honey of infested hives and releasing compounds that function as aggregation pheromones to A. tumida. Since the boron is the toxic element in boric acid, the aim of this article is to assess the amount of boron residues in honey and beeswax from hives treated with the BAA trap. For this aim, the amount of bioaccumulated boron in products of untreated hives was first determined and then compared with the amount of boron of products from hives treated with the BAA trap in two distinct climatic and soil conditions. The study was conducted in the cities of Padilla, Tamaulipas, and Valladolid, Yucatan (Mexico) from August 2014 to March 2015. The quantity of boron in honey was significantly less in Yucatan than in Tamaulipas; this agrees with the boron deficiency among Luvisol and Leptosol soils found in Yucatan compared to the Vertisol soil found in Tamaulipas. In fact, the honey from Yucatan has lower boron levels than those reported in the literature. The BAA treatment was applied for four months, results show that the BAA trap does not have any residual effect in either honey or wax; i.e., there is no significant difference in boron content before and after treatment. On the other hand, the organophosphate pesticide coumaphos was found in 100% of wax samples and in 64% of honey samples collected from Yucatan. The concentration of coumaphos in honey ranges from 0.005 to 0.040 mg/kg, which are below Maximum Residue Limit (MRL) allowed in the European Union (0.1 mg/kg) but 7.14% of samples exceeded the MRL allowed in Canada (0.02 mg/kg).

Discoloration and Adsorption of Acaricides from Beeswax

AbstractThe bleaching effect of solid adsorbent material was evaluated on acaricides residues during the recycling of old wax combs process. The effect of parameters such as initial concentration, moisture level, clay dosing, temperature, particle size, agitation time, Freundlich isotherm and kinetic adsorption were investigated and simulated in pilot plant following industry guidelines. The efficiency capacity of sorbent was assayed in optimal conditions on different levels of chlorfenvinphos, tau‐fluvalinate and coumaphos residues from 0.015 to 88.6 mg/kg, identified in 66 samples of commercial beeswaxes analyzed before and after the process of refination. Experimental results indicated that the use of a dough formed by active carbon (Norit SA4 PAH) and bleaching earth (Tonsil Supreme 114 FF) in the proportion (0.04/4.0 wt % beeswax) under bleaching conditions [agitation (250 rpm) for 45 min working at 90°C under atmospheric pression] was very efficient to reduce color (>80%) and amount of chlorfenvinphos (93.4 ± 7.40%) and coumaphos (93.1 ± 3.90%). But, lower affinity was identified in tau‐fluvalinate pyrethroid (average, 35%) with large variations (13.7–73.9%). The Lagergen pseudo‐first‐order kinetic model was appropriate to organophosphorus acaricides sorption with coefficients of correlation R2 = 0.95 for all initial concentrations studied.Practical ApplicationsBeeswax is a complex mixture of paraffinic hydrocarbons, esters of fatty acids and alcohols, diesters, free fatty acids and alcohols produced by the worker bee. An important aspect is the recycling of beeswax after the extraction of honey. At first, its color is white, but after the bees’ manipulation, it changes from yellow to brownish originated from propolis and pollen pigments. If beeswax is dark its value is much lower. Thus, bleaching is one the most important steps in recycled beeswax. Furthermore, in the last ten years, the accumulated acaricides residues used in the fight against Varroa destructor are clearly detected in commercial beeswaxes limiting its use as an authorised food additive in the EU (code E901). The idea of simultaneous bleaching and decontamination process was introduced in this work to recycle beeswax. The results indicated that the adsorption was very effective in a beeswax melting solution with a simple and economical application.

The presence of synthetic acaricides in beeswax and its influence on the development of resistance in Varroa destructor

After honey production, beeswax ranks second as regards hive product used in industry and cosmetics. In Argentina, the use of commercial wax adulterated with paraffin and other olefins for comb foundation is a common practice. As regards beehives, the progressive accumulation of synthetic acaricides in wax has caused adverse effects on bees, mainly on individuals at different stages of development. Another issue associated with the use of synthetic acaricides is the phenomenon of resistance. This study aimed to determine the presence of adulterants and acaricide residues in commercial wax used in Argentina. Furthermore, the relationship between coumaphos content in wax and the development of mite resistance reported in recent years in Argentina was investigated. The results demonstrate that paraffin is the most common contaminant substance present in recycled beeswax and commercial wax used for comb foundation in the country. Coumaphos was also found to be the most common acaricide present in wax; 87% in commercial and 80% in recycled wax. Fluvalinate was detected in 33% of commercial wax samples and in 27% of recycled wax. A relationship between coumaphos residues and resistance was also established. Future studies should be conducted to establish the mechanisms by which the buildup of acaricides in the beeswax affects the development of resistance in populations of varroa.

The Potential for Using Ozone to Decrease Pesticide Residues in Honey Bee Comb

Ozone is a strong oxidizer, and we evaluated its potential to eliminate pesticides from honeycomb and empty honey bee hives. Honey bees are exposed to pesticides when foraging for nectar and pollen and when beekeepers use in-hive chemical pest control measures. Persistent pesticides can accumulate in the hive over years, potentially harming the bees. Honeycomb is removed from bee colonies for honey extraction and then placed back on the colonies at a later date, providing a time when combs could be fumigated to eliminate or reduce pesticide residues. We found that ozone gas at a rate >920 mg O3/m3 for 10-20 h lowers coumaphos residues on a glass surface by 93-100% and tau-fluvalinate by 75-98%. Ozone was less effective at eliminating pesticides on beeswax, and residues were more effectively eliminated with new combs (comb built by bees within 3 y) than with old combs (combs used by beekeepers for >10 y). Ozone significantly reduced dimethylphenyl formamide, chlorpyrifos, and fenpyroximate contaminations in comb. When comb is treated with ozone, an off-odor is created, but the volatiles were found to be primarily straight chain aldehydes and carboxylic acids that are probably harmless to bees and humans. Ozone may have some utility for lowering pesticide residues in bee hives, but it would be more effective if a mechanism could be found that provides better penetration into wax, a goal not fully accomplished in our method.

Exposure to Pesticides at Sublethal Level and Their Distribution Within a Honey Bee (Apis mellifera) Colony

Honey bee colonies were exposed to pesticides used in agriculture or within bee hives by beekeepers: coumaphos; diazinon; amitraz or fluvalinate. Samples of bee workers, larvae and royal jelly were analysed using Gas Chromatography-Electron Capture Detection (GC-ECD). Amitraz was quantified using High Performance Liquid Chromatography (HPLC), and Gas Chromatography-Tandem Mass Spectrometry (GC/MS/MS) was used for quantification of diazinon. Sixth day after treatment, coumaphos was found in the royal jelly (250 ng/g) secreted by nurse workers and fluvalinate was found in both bee heads (105 ng/g, 8 days after treatment) and in larvae (110 ng/g, 4 days after treatment). Amitraz residues in all sampled material were below the level of detection of 10 ng/g. Diazinon was not detected in any of the analysed samples. The large quantities of fluvalinate found in bee heads and larvae, the coumaphos residues in royal jelly, and additional potential sub-lethal effects on individual honey bees or brood are discussed.

Impact of the Use of Fluvalinate on Different Types of Beeswax from Spanish Hives

Acaricides are applied in agriculture as phytosanitary products against pests and in apiculture to control the bee parasite Varroa destructor. Poor apicultural practices could result in an accumulation of residues in honeybees, in the environment, and in beeswax and other bee products by migration from the wax comb into stored honey through a process of diffusion and consequently constitute a potential risk for humans. In this study, six different types of beeswax samples were analysed for the determination of residues of fluvalinate, coumaphos, and bromopropylate and its metabolite 4,4'-dibromobenzophenone, all of which are the most commonly acaricides used by Spanish beekeepers against V. destructor. The analytic method consists of solid-phase extraction on a SPE Florisil cartridge and high-performance liquid chromatography separation using a photo diode array detector. The results show that fluvalinate residues were detected in 36.3% of samples, ranging from 1.2 to 6.6 microg/g wax. Residues of coumaphos, bromopropylate, and 4,4'-dibromobenzophenone were not found to be greater than their detection limits. This study indicates that the analysis of these compounds in beeswax samples could be used as bioindicators of fluvalinate sanitary treatment and handling practices applied by beekeepers.

Validation of GC-MS method for determination of varroacide residues in propolis

Propolis is a resinous material collected by honeybees (Apis mellifera L.) from various plants and enriched with bee salivary gland secretions and wax. Number of factors, such as beekeeping practice, can affect the quality and consequently the therapeutic value of propolis and its preparations. The most important source of propolis contamination is improper administration of varroacides that have to be used for long-term control of honeybee mite Varroa destructor. Therefore, the analysis of these noxious substances is essential in quality control of propolis. In this study the method based on the extraction with n-hexane followed by purifying of extract on florisil column was used prior to sample analysis. Purified extracts were analyzed by gas chromatography-mass spectrometry hyphenated technique. The procedure was optimized and validated for the determination of bromopropylate, amitraz and coumaphos residues in propolis. Investigated validation parameters of applied procedure were selectivity, linearity, precision, accuracy, robustness and limits of detection (LOD) and quantification (LOQ). All these parameters satisfied the ICH criteria [1] in the test samples of standards mixture for all three examined compounds. The applied method for varroacide analysis in validation sample of propolis was suitable for determination of bromopropylate and coumaphos. Recoveries were 100% for bromopropylate and 63% for coumaphos with LOQ of 0.08 and 1.5µg/g, respectively. On the other hand, the procedure was inappropriate for determination of amitraz, probably due to its instability in complex matrix of propolis as was previously described for honey and beeswax [2].

Miticide Residues in Virginia Honeys

Fifty honey samples from Virginia USA were analyzed for the presence of fluvalinate and coumaphos residues. Samples were collected from hives and from bottled honey provided by beekeepers. No coumaphos or fluvalinate residues above the limit of quantification (0.05 mg/kg) were detected in any of the samples, although trace levels (<0.05 mg/kg) of coumaphos were detected in three samples from hives and trace levels of fluvalinate were found in one hive sample. No residues were detected in any of the bottled honey samples and none of the samples exceeded the US EPA tolerance levels for either miticide.

Bee Honey as an Environmental Bioindicator of Pesticides’ Occurrence in Six Agricultural Areas of Greece

The pollution of six agricultural areas of Greece (north, central, south) by insecticides used in crop protection has been investigated utilizing, as a bioindicator, bee honey produced in those areas. Honey samples collected randomly from apiaries located in those areas were analyzed for pesticide residues with a multianalytical method, able to determine simultaneously up to 10 organophosphorous insecticides from the same honey extract. Findings concerning the acaricide coumaphos were also included, even though it is not used in crop protection. Coumaphos is used to control the mite Varroa destructor, an external parasite of the honeybee. The above areas are cultivated in large extent with citrus trees or cotton or sunflower crops, which are good forages for honeybees. The main pests of those crops are insects; hence, insecticides are used on a large scale for crop protection. The most contaminated samples originated from citrus groves; 16 out of 19 had pesticide residues: 4 samples had chlorfenvinphos (21.05%), 10 had chlorpyrifos (52.63%) and 2 had phorate (10.53%). Out of 17 samples from cotton fields, residues were found in 8, phorate in 6 (35.29%), chlorfenvinphos in 1 (5.88%), and chlorpyrifos in 1 (5.88%). Out of nine samples from fields of sunflower, four had phorate residues (44.44%). In brief, from the 50 analyzed samples, residues of chlorfenvinphos were detected in 5 samples (10%), residues of chlorpyrifos in 11 samples (22%), and residues of phorate in 12 samples (24%). Their levels ranged between 0.70 and 0.89 microg/kg. Coumaphos residues ranged from 0.10 up to 4.80 microg/kg and were derived exclusively from beehives treated with Perizin (the commercial formulation of coumaphos) for Varroa control. This study indicates that in agricultural areas with developed apiculture, useful information about the occurrence and the distribution of pesticide residues due to crop protection treatments can be derived from the analysis of randomly collected honey samples, used as bioindicators. It also shows that, very often, the chemicals used by apiculturists inside the hives in order to control disease are the main pollutants of the produced honey.

Residue distribution of the acaricide coumaphos in honey following application of a new slow‐release formulation

Acaricide used in beehives for the control of varroa often leaves residues in bee products. The behaviour and distribution of the acaricide coumaphos in honey following the application of a new slow-release strip formulation (CheckMite+) was assessed. The bee colonies were allowed to build new combs without foundation, and two strips were hung in the brood chamber of each colony for a period of 42 days. The distribution of coumaphos residues in honey in relation to the position of the frame and the duration of treatment was examined by collecting samples from each comb at various time intervals up to 145 days after treatment. In the brood chamber, coumaphos was incorporated into honey from the first day of application, and residues accumulated mainly in combs placed next to strips. In the adjacent combs, residues remained at low concentrations with slight variations. In the honey chamber, residue concentrations on the day of strip removal ranged between 0.006 and 0.020 mg kg(-1), while 79 days after application the concentration of coumaphos residues was below 0.020 mg kg(-1). Residues above the EC fixed maximum residue limit (MRL) of 0.1 mg kg(-1) were measured only in brood chamber honey obtained from those combs placed next to strips. In these samples, 0.060-0.111 mg kg(-1) of coumaphos was detected up to 103 days after strip removal. Coumaphos residues in honey extracted from combs that were placed at the edge of the brood chamber were found below the MRL value, even during the 42 day period of CheckMite+ strip treatment.

Acaricide residues in honey and wax after treatment of honey bee colonies with Apivar®or Asuntol®50

Acaricide residues were assessed in French commercial beeswax using newly developed liquid and gas chromatography methods. Most of the commercial wax samples and all samples taken during the industrial recycling process contained coumaphos and fluvalinate. Amitraz and coumaphos residue levels were also followed in several hives experimentally treated with Asuntol®50 or Apivar®, two products used in France to control varroa infestation. After the Asuntol®50 treatment, coumaphos residues increased in honey and wax combs, persisted more than 30 days in honey and one year or more in comb wax. The half-life of coumaphos was 69 and 115–346 days in honey and comb wax respectively. Following Apivar® treatment, amitraz was not detected in honey nor in wax. These results are consistent with and complete other studies: the use of coumaphos entails wax contamination which persists through commercial recycling. As this may be a threat for bee health, the use of Asuntol®50 should be avoided.

Determination of Acaricide Residues in Saudi Arabian Honey and Beeswax Using Solid Phase Extraction and Gas Chromatography

Determination of acaricide residues of flumethrin, tau-fluvalinate, coumaphos, and amitraz in honey and beeswax was carried out using a rapid extraction method utilizing C-18 SPE cartridges and an analytical method utilizing GC with ECD, NPD, and MSD detectors for the four acaricides. Recovery percentages from the extraction method ranged from 90–102%, while the minimum detection levels ranged from 0.01–0.05 mg/kg for the acaricides. Nine of the 21 analyzed samples were found to be contaminated with the acaricides tau-fluvalinate and coumaphos. Neither flumethrin nor amitraz was detected in any of the honey or wax samples. Coumaphos was found only in honey samples in which two samples exceeded the tolerance levels set by EPA and EC regulations. It has not been detected in beeswax. Five honey samples and eight beeswax samples were found to be contaminated with tau-fluvalinate. One of the wax samples was contaminated with a relatively high residue of tau-fluvalinate and contained above 10 mg/kg.

Analysis of organophosphorus pesticides in honeybee by liquid chromatography-atmospheric pressure chemical ionization-mass spectrometry.

Pesticides applied in extended agricultural fields may be controlled by means of bioindicators, such as honeybees, in which are the pesticides bioaccumulate. Liquid chromatography-atmospheric pressure chemical ionization-mass spectrometry (LC-APCI-MS) experiments with positive (PI) and negative (NI) ion modes were optimized for the analysis of 22 organophosphorus pesticides in honeybee samples. The extraction required 3 g of sample, which was extracted with acetone. The extract was purified with coagulating solution and reextracted with Cl(2)CH(2). Pesticides studied could be detected by both ionization modes except for parathion, parathion-methyl, and bromophos, which did not give signals in PI mode, and triazophos, which was not detected in NI mode. Fragmentation voltage and vaporizer temperature were optimized to achieve the highest sensitivity. The spectra profile of each pesticide in PI mode showed the [M + H](+) ion as the main signal, whereas in NI mode only fragment ions were shown. The detection limit obtained in selected ion monitoring mode ranged from 1 to 15 microg kg(-1). The average recoveries from spiked honeybees at various concentration levels (0.5-5 mg kg(-1)) exceeded 65% with relative standard deviations of 4-15%. The method was applied to real samples, in which residues of coumaphos and dimethoate were detected.