Acaricide resistance status of deltamethrin and coumaphos in Hyalomma anatolicum ticks collected from different districts of Haryana.

To study the acaricide resistance status and possible mechanisms of action in conferring resistance to commonly used acaricides (deltamethrin and coumaphos), Hyalomma anatolicum ticks were collected from 6 dairy farms of Hisar and Charkhi Dadri districts of Haryana. By using standard larval packet test, H. anatolicum tick larvae of Charkhi Dadri isolates were found to be susceptible (100% mortality) to both the acaricides. Level-I resistance against coumaphos was recorded from four isolates, whereas, level-II was observed in only one isolate, collected from Hisar. One isolates (Kaimri) from Hisar also showed level-I resistance against deltamethrin. Biochemically, the ticks having higher values of resistance factor (RF) against coumaphos were found to possess increased enzymatic activity of α-esterase, ß-esterase, glutathione-S-transferase (GST) and mono-oxygenase enzymes, whereas, the monoamine oxidase did not show any constant trend. However, the RF showed a statistical significant correlation with GST only. Native PAGE analysis of H. anatolicum ticks revealed the presence of nine types of esterases (EST-1 h to EST-9 h) by using napthyl acetate as substrate. In the inhibitory assay, esterases were found to be inhibited by PMSF, indicating the presence of serine residue at catalytic triad. The partial cds of carboxylesterase and domain II of sodium channel genes were sequenced to determine any proposed mutations in resistant isolates of H. anatolicum ticks, however, no mutations were observed in either gene, indicating that increased expression of detoxification enzymes as a possible mechanism for resistance development, in the current study.

Status of Acaricide Resistance and Detecting the Knockdown Resistance Mutation T2134A in the Cattle Tick Rhipicephalus microplus (Acari: Ixodidae) from Northeastern Mexico.

Rhipicephalus microplus is the most important tick in veterinary medicine, given its repercussions on animal production. The principal strategy to avoid adverse effects associated with R. microplus is the chemical control of tick populations through organosynthetic acaricides. Therefore, monitoring susceptibility to acaricides is paramount in any control program. This study aimed to analyze the resistance status of 2 populations of R. microplus from northeastern Mexico to the organochlorine (OC) lindane, organophosphates (OP) coumaphos, chlorfenvinphos, diazinon, and chlorpyrifos, and the synthetic pyrethroids (SPs) flumethrin, deltamethrin, and cypermethrin. Discriminating doses (DD) of each acaricide were used in the larval packet bioassay (LPT). Additionally, the presence of the knockdown resistance (kdr) mutation T2134A associated with pyrethroid resistance was evaluated using allele-specific polymerase chain reaction (PCR). The populations of R. microplus showed a high frequency of resistance to SP, with mortality rates of less than 5%; they also showed resistance to the OPs (diazinon and chlorpyrifos) with mortality rates ranging from 1.29% to 34.62%; meanwhile, they were susceptible to coumaphos and chlorfenvinphos. Mortality rates higher than 66% were observed for lindane, indicating susceptibility. The mutant allele of the kdr mutation T2134A was detected in 75% and 100% of the pools analyzed. The populations studied presented a highly resistant profile to pyrethroids, with the presence of the kdr mutant allele A2134. The susceptibility to the organophosphates such as coumaphos and chlorfenvinphos of R. microplus from northeastern Mexico should be noted.

Histometric and morphological damage caused by Serratia marcescens to the tick Rhipicephalus microplus (Acari: Ixodidae).

Rhipicephalus microplus tick is the ectoparasite causing the greatest economic losses in the livestock industry. Multi-resistance in ticks is increasing, generating the inefficiency of traditional ixodicides, for which biological control has been proposed as an alternative. In this work, we analyze the histomorphological damage caused by the bacterial strain EC-35 on Rhipicephalus microplus. The ixodicidal effect of EC-35 total protein was evaluated on larval or adult ticks comparing with the commercial ixodicide coumaphos 0.02% as a control. Female ticks were processed using the paraffin-embedding technique and stained with hematoxylin-eosin. Also, the pathogenicity of EC-35 was evaluated by capillary feeding and coelom inoculation tests. The identification of the bacterium was performed using the molecular markers 16S RNA and rpoB, by PCR and sequencing technique, and the evolutionary distance was analyzed by Bayesian phylogenetic inference. No differences were observed in the perimeter and area of larvae treated with EC-35 or Coumaphos. The thickness of the integument decreased a 65% with the EC-35 treatment (6.01 ± 0.6 µm) and of 30% in coumaphos (12.04 ± 1.2 µm) in larvae compared with the control group (18.41 ± 2 µm), while no difference was found in adult ticks. The capillary feeding test and coelom inoculation with EC-35 showed an inhibition of reproductive potential of 99.8 ± 7 and an oviposition Inhibition 97 ± 3.02%. The EC-35 strain was genetically related to Serratia marcescens, concluding that these bacteria caused high mortality, oviposition Inhibition, and integument thinning and drastic loss of histoarchitecture in R. microplus tick larvae.

Characterization of deltamethrin, cypermethrin, coumaphos and ivermectin resistance in populations of Rhipicephalus microplus in India and efficacy of an antitick natural formulation prepared from Ageratum conyzoides.

Rhipicephalus microplus is posing a serious threat to productive animal husbandry. Excessive use of synthetic chemicals in tick management has led to the development of resistant tick populations. Characterization of resistance to deltamethrin, cypermethrin, coumaphos and ivermectin in ticks is necessary to develop a suitable and sustainable control strategy. Based on adult immersion test and larval packet test, the resistance ratios (RR50) for adults and larvae of R. microplus populations from two Indian states ranged from 3.8 to 19.4 and 1.35-25.0 against deltamethrin, 0.061-26.3 and 0.22-19.2 against cypermethrin, and 0.2-9.5 and 0.01-3.1 against coumaphos, respectively, were recorded. Moreover, the RR50 for adults ranged from 0.212 to 3.87 against ivermectin. The RR50 for different acaricides was significantly (p<0.01) correlated with esterases, Glutathione S-transferase and monooxygenase activity. A point mutation at the 190th position of the domain II S4-5 linker region of the sodium channel gene in synthetic pyrethroids (SP) resistant populations was also detected. An antitick natural formulation prepared from the plant Azeratum conyzoides and containing two major compounds, Precocene-I (7­methoxy-2, 2-dimethyl 2H-chromene) and Precocene II (6, 7-dimethoxy-2, 2-dimethyl- 3-chromene), was developed and tested against the resistant ticks. The LC50 values of the natural formulation against the resistant populations were in the range of 4.31-5.33% irrespective of their RR50 values. Multi-acaricide resistant populations of R. microplus are established in India and the A. conyzoides based natural formulation can be used for its management.

Reduced proinsecticide activation by cytochrome P450 confers coumaphos resistance in the major bee parasite Varroa destructor.

Varroa destructor is one of the main problems in modern beekeeping. Highly selective acaricides with low toxicity to bees are used internationally to control this mite. One of the key acaricides is the organophosphorus (OP) proinsecticide coumaphos, that becomes toxic after enzymatic activation inside Varroa We show here that mites from the island Andros (AN-CR) exhibit high levels of coumaphos resistance. Resistance is not mediated by decreased coumaphos uptake, target-site resistance, or increased detoxification. Reduced proinsecticide activation by a cytochrome P450 enzyme was the main resistance mechanism, a powerful and rarely encountered evolutionary solution to insecticide selection pressure. After treatment with sublethal doses of [14C] coumaphos, susceptible mite extracts had substantial amounts of coroxon, the activated metabolite of coumaphos, while resistant mites had only trace amounts. This indicates a suppression of the P450 (CYP)-mediated activation step in the AN-CR mites. Bioassays with coroxon to bypass the activation step showed that resistance was dramatically reduced. There are 26 CYPs present in the V. destructor genome. Transcriptome analysis revealed overexpression in resistant mites of CYP4DP24 and underexpression of CYP3012A6 and CYP4EP4 RNA interference of CYP4EP4 in the susceptible population, to mimic underexpression seen in the resistant mites, prevented coumaphos activation and decreased coumaphos toxicity.

Assessing the resistance to acaricides in Varroa destructor from several Spanish locations.

Varroosis is the disease caused by the ectoparasitic mite Varroa destructor, one of the most destructive diseases of honeybees. In Spain, there is great concern because there are many therapeutic failures after acaricide treatments intended to control varroosis outbreaks. In some of these cases it is not clear whether such failures are due to the evolution of resistance. Therefore, it is of high interest the development of methodologies to test the level of resistance in mite populations. In this work, a simple bioassay methodology was used to test whether some reports on low efficacy in different regions of Spain were in fact related to reduced Varroa sensitivity to the most used acaricides. This bioassay proved to be very effective in evaluating the presence of mites that survive after being exposed to acaricides. In the samples tested, the mortality caused by coumaphos ranged from 2 to 89%; for tau-fluvalinate, it ranged from 5 to 96%. On the other hand, amitraz caused 100% mortality in all cases. These results suggest the presence of Varroa resistant to coumaphos and fluvalinate in most of the apiaries sampled, even in those where these active ingredients were not used in the last years. The bioassay technique presented here, either alone or in combination with other molecular tools, could be useful in detecting mite populations with different sensitivity to acaricides, which is of vital interest in selecting the best management and/or acaricide strategy to control the parasite in apiaries.

Comparative impact of coumaphos, amitraz and plant extract of Ageratum conyzoides on the oogenesis of Rhipicephalus microplus.

The present experiment was conducted to evaluate and compare the impact of Ageratum conyzoides plant extract (ACE) with routinely used synthetic acaricides i.e., amitraz and coumaphos on the oogenesis of engorged adult females of Rhipicephalus microplus tick. On the day of dropping from the host, panoistic ovary of R. microplus appeared white in colour, horseshoe shaped, hollow tubular organ with immature oocytes predominantly in dorsal groove. Different developmental stages of oocytes (I-V) proceed simultaneously and asynchronously. Oocytes showed gradual increase in size, deep brown colored with accumulation of eggs in oviduct during 24-72 hours of development.At LC90 concentration a highly significant (p < 0.001) cessation of egg laying after exposure to amitraz and ACE while significant reduction (p < 0.01) of egg laying in coumaphos treated ticks was observed. Upon dissection of treated ticks, uterus and oviduct packed with eggs, which failed to pass out was observed. The histo-architectural alterations including presence of extensive vacuolation, alteration of oocyte morphology, deformation of chorion and disorganization of yolk granules were observed in the treated ovaries. Histochemically, low level of storage or synthesis of essential elements viz., proteins, polysaccharides and lipids in treated oocytes responsible for reduction of fertility and inhibition of progress of vitellogenesis was observed.

Heat shock proteins in Varroa destructor exposed to heat stress and in-hive acaricides.

Varroa destructor is one of the major pests that affect honeybees around the world. Chemical treatments are common to control varroosis, but mites possess biochemical adaptive mechanisms to resist these treatments, enabling them to survive. So far, no information is available regarding whether these pesticides can induce the expression of heat shock protein (Hsp) as a common protective mechanism against tissue damage. The aims of this study were to determine differences in heat shock tolerance between mites collected from brood combs and phoretic ones, and to examine patterns of protein expression of Hsp70 that occur in various populations of V. destructor after exposure to acaricides commonly employed in beekeeping, such as flumethrin, tau-fluvalinate and coumaphos. Curiously, mites obtained from brood cells were alive at 40 °C, unlike phoretic mites that reached 100% mortality, demonstrating differential thermo-tolerance. Heat treatment induced Hsp70 in mites 4 × more than in control mites and no differences in response were observed in phoretic versus cell-brood-obtained mites. Dose-response assays were carried out at increasing acaricide concentrations. Each population showed a different stress response to acaricides despite belonging to the same geographic region. In one of them, coumaphos acted as a hormetic stressor. Pyrethroids also induced Hsp70, but mite population seemed sensitive to this treatment. We concluded that Hsp70 could represent a robust biomarker for measuring exposure of V. destructor to thermal and chemical stress, depending on the acaricide class and interpopulation variability. This is relevant because it is the first time that stress response is analyzed in this biological model, providing new insight in host-parasite-xenobiotic interaction.

Prochloraz and coumaphos induce different gene expression patterns in three developmental stages of the Carniolan honey bee (Apis mellifera carnica Pollmann).

The Carniolan honey bee, Apis mellifera carnica, is a Slovenian autochthonous subspecies of honey bee. In recent years, the country has recorded an annual loss of bee colonies through mortality of up to 35%. One possible reason for such high mortality could be the exposure of honey bees to xenobiotic residues that have been found in honey bee and beehive products. Acaricides are applied by beekeepers to control varroosis, while the most abundant common agricultural chemicals found in honey bee and beehive products are fungicides, which may enter the system when applied to nearby flowering crops and fruit plants. Acaricides and fungicides are not intrinsically highly toxic to bees but their action in combination might lead to higher honey bee sensitivity or mortality. In the present study we investigated the molecular immune response of honey bee workers at different developmental stages (prepupa, white-eyed pupa, adult) exposed to the acaricide coumaphos and the fungicide prochloraz individually and in combination. Expression of 17 immune-related genes was examined by quantitative RT-PCR. In treated prepupae downregulation of most immune-related genes was observed in all treatments, while in adults upregulation of most of the genes was recorded. Our study shows for the first time that negative impacts of prochloraz and a combination of coumaphos and prochloraz differ among the different developmental stages of honey bees. The main effect of the xenobiotic combination was found to be upregulation of the antimicrobial peptide genes abaecin and defensin-1 in adult honey bees. Changes in immune-related gene expression could result in depressed immunity of honey bees and their increased susceptibility to various pathogens.

Determination and establishment of discriminating concentrations of malathion, coumaphos, fenvalerate and fipronil for monitoring acaricide resistance in ticks infesting animals.

Discriminating concentrations (DCs) of malathion, coumaphos, fenvalerate and fipronil were determined to monitor acaricide resistance in field conditions. The LC99 values with 95% confidence interval for malathion, coumaphos, fenvalerate and fipronil were 5126.8 (5011.5-5240.7), 131.0 (120.4-142.5), 2257.5 (2198.1-2318.4) and 6.2 (5.87-6.55), respectively. The narrow confidence intervals in LC50 and LC99 of adult immersion test (AIT) and larval packet test (LPT) affirming the homogeneity of IVRI-I line. Variation in LPT based LC50 and LC99 values of malathion (55.9ppm) and coumpahos (28.4ppm) compared to those obtained in AIT indicating that larvae were more susceptible to these chemicals. The DCs for malathion, coumaphos, fenvalerate and fipronil against adults were determined as 10253.6, 262.0, 4515.0 and 12.4ppm while against larvae the values were 111.8, 56.8, 4014.0 and 9.6ppm, respectively. The working efficiency of DCs was successfully tested in field tick isolates. Establishment of country specific DCs of commonly used insecticides for monitoring of resistance in field ticks is emphasized for establishing tick control strategies.

Cholinergic pesticides cause mushroom body neuronal inactivation in honeybees.

Pesticides that target cholinergic neurotransmission are highly effective, but their use has been implicated in insect pollinator population decline. Honeybees are exposed to two widely used classes of cholinergic pesticide: neonicotinoids (nicotinic receptor agonists) and organophosphate miticides (acetylcholinesterase inhibitors). Although sublethal levels of neonicotinoids are known to disrupt honeybee learning and behaviour, the neurophysiological basis of these effects has not been shown. Here, using recordings from mushroom body Kenyon cells in acutely isolated honeybee brain, we show that the neonicotinoids imidacloprid and clothianidin, and the organophosphate miticide coumaphos oxon, cause a depolarization-block of neuronal firing and inhibit nicotinic responses. These effects are observed at concentrations that are encountered by foraging honeybees and within the hive, and are additive with combined application. Our findings demonstrate a neuronal mechanism that may account for the cognitive impairments caused by neonicotinoids, and predict that exposure to multiple pesticides that target cholinergic signalling will cause enhanced toxicity to pollinators.

Organophosphorus hydrolase: a multifaceted plant genetic marker which is selectable, scorable, and quantifiable in whole seed.

Organophosphorus hydrolase (OPH, EC 3.1.8.1) provides a novel function as an alternative genetic marker system for use in many types of plant transformations. OPH is a high-capacity hydrolase with multiple organophosphorus substrates, many of which are neurotoxins and thus used extensively as pesticides. This spectrum of organophosphates includes compounds that are phytotoxic as well as those that are hydrolyzed to products that are easily detected visually without significant disruption of plant health. This dichotomy gives OPH the features of both a selectable marker as well as that of a scorable marker system, and these characteristics have been tested at several stages during the plant transformation and regeneration process. Finally, it is possible to quantify hydrolytic activity in the seed without interfering with its subsequent growth and regeneration.

Susceptibility of Varroa destructor (Acari: Varroidae) to synthetic acaricides in Uruguay: Varroa mites' potential to develop acaricide resistance.

The purpose of this study was to estimate the acaricide susceptibility of Varroa destructor populations from Uruguay, which had never been exposed to synthetic acaricides. It was also to determine whether acaricide resistance to coumaphos occurred in apiaries in which acaricide rotation had been applied. Bioassays with acaricides against mite populations that had never been exposed to synthetic acaricides were performed, also against mite populations in which control failures with coumaphos had been reported. Additionally, coumaphos' effectiveness in honeybee colonies was experimentally tested. The lethal concentration that kills 50% of the exposed animals (LC(50)) for susceptible mite populations amounted to 0.15 µg/Petri dish for coumaphos and to less than 0.3 µg/Petri dish for the other acaricides. Coumaphos LC(50) was above 40 µg/Petri dish for resistant mites. The effectiveness of coumaphos in honeybee colonies parasitized by V. destructor ranged from 17.6% to 93.9%. LC(50) for mite populations susceptible to the most commonly applied miticides was determined, and the first case of coumaphos resistance recorded in Uruguay was established.

Synergistic interactions between in-hive miticides in Apis mellifera.

The varroa mite, Varroa destructor Anderson & Trueman, is a devastating pest of honey bees, Apis mellifera L., that has been primarily controlled over the last 15 yr with two in-hive miticides: the organophosphate coumaphos (Checkmite+), and the pyrethroid tau-fluvalinate (Apistan). Both coumaphos and tau-fluvalinate are lipophilic compounds that are absorbed by the wax component of the hive, where they are stable and have the potential to build up over repeated treatments such that bees could be exposed to both compounds simultaneously. Although these compounds were chosen as in-hive miticides due to their low toxicity to honey bees, that low toxicity depends, at least in part, on rapid detoxification mediated by cytochrome P450 monooxygenase enzymes (P450s). In this laboratory study, we observed a large increase in the toxicity of tau-fluvalinate to 3-d-old bees that had been treated previously with coumaphos, and a moderate increase in the toxicity of coumpahos in bees treated previously with tau-fluvalinate. The observed synergism may result from competition between miticides for access to detoxicative P450s. These results suggest that honey bee mortality may occur with the application of otherwise sublethal doses of miticide when tau-fluvalinate and coumaphos are simultaneously present in the hive.

Transient expression of organophosphorus hydrolase to enhance the degrading activity of tomato fruit on coumaphos.

We constructed an expression cassette of the organophosphorus pesticide degrading (opd) gene under the control of the E8 promoter. Then opd was transformed into tomato fruit using an agroinfiltration transient expression system. beta-Glucuronidase (GUS) staining, reverse transcription-polymerase chain reaction (RT-PCR), wavelength scanning, and fluorescent reaction were performed to examine the expression of the opd gene and the hydrolysis activity on coumaphos of organophosphorus hydrolase (OPH) in tomato fruit. The results show that the agroinfiltrated tomato fruit-expressed OPH had the maximum hydrolysis activity of about 11.59 U/mg total soluble protein. These results will allow us to focus on breeding transgenic plants that could not only enhance the degrading capability of fruit and but also hold no negative effects on pest control when spraying organophosphorus pesticides onto the seedlings in fields.

Differential response to diazinon and coumaphos in a strain of Boophilus microplus (Acari: Ixodidae) collected in Mexico.

Boophilus microplus, collected from Nuevo Leon, Mexico, were found to be highly resistant to diazinon but not highly resistant to coumaphos, suggesting that different mechanisms of resistance were present in these ticks than other Mexican organophosphate (OP)-resistant ticks reported previously. When exposed to coumaphos and piperonyl butoxide or triphenylphosphate, the LCso estimate was reduced by 3.5- and 6.3-fold, respectively, suggesting that mono-oxygenases and/or esterases were involved in resistance to coumaphos. Additionally, it was determined that this strain had an Acetylycholinesterase (AChe) that was insensitive to the active form of coumaphos, coroxon, taking at least 24 min longer to reach 50% reduction in AChE activity compared with the susceptible strain. When exposed to diazinon, none of the synergists tested significantly lowered the LC50. However, it was determined that it took six times longer to reach 60% inhibition of AChE in the resistant strain compared with the susceptible strain when exposed to the active form of diazinon, diazoxon. Insensitive AChE seems to be very common in OP-resistant B. microplus. The potential benefits for the development of a field-portable AChE inhibition assay kit are discussed.

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.

Effect of coumaphos on cholinesterase activity, hematology, and biochemical blood parameters of bovines in tropical regions of Mexico.

To assess the effect of coumaphos [O-(3-chloro-4-methyl-2-oxo-2H-1-benzopyran-7-yl) O,O-diethyl phosphorothioate] exposure on physiological responses during bovine production, acetylcolinesterase (AChE) and butyrylcholinesterase (BuChE) activities were measured in whole blood, erythrocytes, and plasma of healthy male steers (Bos Taurus x Bos indicus) sprayed with coumaphos at a non-lethal dose of 1 mg kg(- 1) body weight per day once every 14 (in vivo group) or 21 days (southern and central groups). Coumaphos topically administered at 1 mg/kg body weight per day to cattle under normal management practices in tropical areas produced a significant inhibition in erythrocyte (RBC) AChE and BuAChE activities when compared to baseline levels. RBC-AChE activity for the in vivo group decreased 71.3% (P < 0.05) and BuChE activity 59.1% (P < 0.05); RBC-AChE activity decreased 55.1% (P < 0.05) (southern group) and 43.4% (P < 0.05) (central group). Compared to the control specimens, steers from in vivo, southern, and central groups after 150 days of exposure had lower (P < 0.05) leukocyte count, absolute lymphocyte, erythrocyte, and platelet counts. Decreases in RBC-AChE activities correlated with decreased lymphocyte (r = 1.000, p = 0.01), erythrocyte (r = 1.000, p = 0.003), and platelet counts (r = 0.841, p = 0.036). Significantly increased BUN levels (P < 0.05) correlated with the decrease in RBC-AChE activities (r = - 0.997, p = 0.047) and with the decrease in absolute red blood cell (r = - 0.883, p = 0.020) and lymphocyte (r = - 0.825, p = 0.043) counts; increased (P < 0.05) total plasma protein levels correlated with the decrease in RBC-AChE activities (r = -0.998, p = 0.043), absolute red blood cell (r = - 0.998, p = 0.040), lymphocyte (r = - 0.893, p = 0.017), and platelet (r = -0.855, p = 0.030) counts. The physiological responses correlated with the erythrocyte acetylcholinesterase inhibition could be considered as early indicators or warning responses of bovine exposures to organophosphorus pesticides (OPs).

Serial analysis of gene expression in the southern cattle tick following acaricide treatment of larvae from organophosphate resistant and susceptible strains.

Organophosphate resistant and susceptible tick larvae from laboratory strains of the southern cattle tick, Rhipicephalus (Boophilus) microplus were exposed to low doses of the organophosphate (OP) acaricide, coumaphos. Serial analysis of gene expression (SAGE) was used to analyse differential gene expression in response to OP treatment and to compare the responses of OP-treated and -untreated resistant and susceptible tick larvae. An R. microplus Gene Index was used as an EST database to identify genes which corresponded to SAGE tags whose abundance changed in response to acaricide exposure. Relative quantitative RT-PCR was used to confirm the differential expression results from the SAGE experiments. Of particular interest is a SAGE tag which corresponds to a cytochrome P450-like EST in the Gene Index which was more abundant in untreated OP resistant larvae compared to untreated OP susceptible larvae. This SAGE tag was also more abundant in OP resistant larvae treated with OP compared to OP susceptible larvae treated with OP.

Formic acid treatment for control of Varroa destructor (Mesostigmata: Varroidae) and safety to Apis mellifera (Hymenoptera: Apidae) under southern United States conditions.

The efficacy of a formic acid pad formulation was field tested for control of the honey bee parasitic mite Varroa destructor Anderson & Trueman in Florida and Texas. This pad formulation gave 39.8 +/- 11.1% control at the end of a 6-wk treatment period, which did not significantly differ from the initial sample date. Coumaphos treatment provided poor control (38.4 +/- 11.1%) over the 6-wk period, confirming reports of coumaphos resistance in the region. Under relatively warm winter conditions in southern Texas, formic acid caused mortality of developing eggs and brood. If resistance by V. destructor to the two acaricides registered for its control in the United States continues, the formic acid pad could provide an alternative compound to use as part of an integrated pest management approach. Given the low control seen in this trial, however, modifications of application technology would seem necessary.