Ulrich R. Bernier

Elution patterns from capillary GC for methyl-branched alkanes

A common and confusing problem in analyses of insect hydrocarbons is in making sense of complicated gas chromatograms and interpreting mass spectra since branched chain compounds differing by one or two carbons in backbone or chain length may elute from the column at nearly the same time. To address this confusing situation, relative gas chromatography (GC) retention times are presented for typical mono-, di-, tri-, and tetramethylalkanes comprising most of the commonly appearing series of homologous methyl-branched alkanes up to 53 carbons that are found in insect cuticular hydrocarbons. Typical insect-derived methylalkanes with backbones of 33 carbons were characterized by Kovats indices (KI); monomethyl alkanes elute between KI 3328 and 3374, dimethylalkanes elute between KI 3340 and 3410, trimethylalkanes elute between KI 3378 and 3437, and tetramethylalkanes elute between KI 3409 and 3459, depending upon the positions of substituents. A protocol is described for identification of methyl-branched hydrocarbons eluted from nonpolar polysiloxane DB-1 capillary GC columns. In this protocol, retention indices (KI values) are assigned to peaks, then the patterns in GC peaks that probably contain homologs are marked to assist subsequent GC-mass spectrometric (GC-MS) interpretation. Use of the KI allows assignment of likely structures and the elimination of others, with demonstrative consistency, as there are no known exceptions. Interpretation of electron ionization mass spectra can then proceed within narrowed structural possibilities without the necessity of chemical ionization GC-MS analysis. Also included are specific examples of insect hydrocarbons that were assembled from 30 years of the literature, and these are intended to help with confirmation of confusing or contradictory structures.

Synergistic Attraction of Aedes aegypti (L.) to Binary Blends of L-Lactic Acid and Acetone, Dichloromethane, or Dimethyl Disulfide

Abstract Kairomones produced by humans provide female anthropophilic mosquitoes with vital cues used in host-seeking for a blood meal. These chemicals are emanated primarily by the skin and provide the mosquitoes a means to orient themselves to humans at a relatively close range. Chemical studies of these emanations have provided new ideas for the formulation of attractant blends. We report mosquito attraction responses for three binary blends and their separate components. The blends are comprised of L-lactic acid plus either acetone, dichloromethane, or dimethyl disulfide. At the emission rates used in our bioassays, these blends synergistically attract laboratory-reared female Aedes aegypti. Carbon dioxide is not a necessary component to yield high levels of attraction with these blends. It is postulated that at least one of these synergistic blends (L-lactic acid and acetone) produces mosquito attraction behavior similar to L-lactic acid and CO2.

Comparison of Contact and Spatial Repellency of Catnip Oil and N,N-Diethyl-3-methylbenzamide (Deet) Against Mosquitoes

Abstract Nepetalactone, the primary component of catnip oil, was compared with the repellent N,N-diethyl-3-methylbenzamide (deet) for its ability to affect the host-seeking ability of Aedes aegypti (L.). A triple cage olfactometer was used to bioassay each substance and to assess its attraction inhibition (spatial repellent) attributes when combined with the following attractants: carbon dioxide, acetone, a blend of l-lactic acid and acetone, and human odors. Repellent tests were conducted with each substance against female Ae. aegypti, Anopheles albimanus Weidemann, and Anopheles quadrimaculatus Say. Catnip oil and deet were both weakly attractive to Ae. aegypti, catnip oil was the better spatial repellent, whereas deet was a more effective contact repellent in tests with all three species of mosquitoes.

Repellency of IR3535, KBR3023, para-menthane-3,8-diol, and deet to black salt marsh mosquitoes (Diptera: Culicidae) in the Everglades National Park.

Abstract IR3535, KBR3023, para-menthane-3,8-diol (PMD), and deet were evaluated in controlled studies with human subjects (n = 5) for repellency to black salt marsh mosquitoes (Ochlerotatus taeniorhynchus Wiedemann), in the Everglades National Park, FL. In tests of 6-h duration, with an average mosquito biting pressure on exposed forearm skin of 19.5 (±13.7) bites per minute, the mean percent repellencies (SE) for IR3535, KBR3023, PMD, and deet was 88.6 (3.2), 97.5 (1.7), 89.2 (2.9), and 94.8 (2.5), respectively. Mean complete protection times (SE) for IR3535, KBR3023, PMD, and deet were 3.0 (1.0), 5.4 (0.6), 3.8 (1.4), and 5.6 (0.5) h, respectively. Untreated (ethanol) controls provided 0% repellency. When mosquito biting rates on the untreated forearm skin of repellent-treated subjects were compared with biting rates on the forearm skin of control subjects, the former were 23%–40% lower early in tests and as much as 22% higher late in tests. These differences cast doubt on the technical merit of test designs comprising evaluation of more than one repellent at a time on the same human subject while underscoring the importance of untreated subjects as negative controls in field repellent bioassays.

Olfactometric Evaluation of Spatial Repellents for Aedes aegypti

Abstract The spatial repellency responses of Aedes aegypti (L.) to deet, dehydrolinalool and linalool were evaluated using a dual port olfactometer. In the absence of human attractant mixture, each of the three chemicals resulted in activation and/or orientation of mosquitoes to the chemical source. Linalool was the most attractive compound. In the presence of human attractant mixture, activation and/or orientation of mosquitoes to each of the three chemicals was reduced. We compared reductions in mosquito responses to each of the three chemicals, in the presence of human attractant mixture, to estimate spatial repellency. As expected, lowest spatial repellency (7.3%) was observed using human attractant alone. Highest spatial repellency (33.6%) was observed using a combination of linalool and dehydrolinalool. Deet did not manifest spatial repellency, whereas linalool and dehydrolinalool alone, and in combination, exhibited spatial repellency.

Synthesis and bioassay of improved mosquito repellents predicted from chemical structure

Mosquito repellency data on acylpiperidines derived from the U.S. Department of Agriculture archives were modeled by using molecular descriptors calculated by CODESSA PRO software. An artificial neural network model was developed for the correlation of these archival results and used to predict the repellent activity of novel compounds of similar structures. A series of 34 promising N-acylpiperidine mosquito repellent candidates (4a–4q′) were synthesized by reactions of acylbenzotriazoles 2a–2p with piperidines 3a–3f. Compounds (4a–4q′) were screened as topically applied mosquito repellents by measuring the duration of repellency after application to cloth patches worn on the arms of human volunteers. Some compounds that were evaluated repelled mosquitoes as much as three times longer than N,N-diethyl-m-toluamide (DEET), the most widely used repellent throughout the world. The newly measured durations of repellency were used to obtain a superior correlation equation relating mosquito repellency to molecular structure.

Laboratory Evaluation of Avian Odors for Mosquito (Diptera: Culicidae) Attraction

Abstract Attraction of Culex quinquefasciatus Say, Culex tarsalis Coquillett, Culex nigripalpus Theobald, and Aedes aegypti (L.) to avian and other host odors was investigated in a dual-port olfactometer. Although attraction to a human arm was high for Ae. aegypti (>80%) and low for all Culex spp. (<25%), all species responded similarly to a chicken (55.3–73.6%). Responses of Ae. aegypti, Cx. quinquefasciatus, and Cx. nigripalpus to feathers were low (<20%) but greater than to controls. There was no difference in attraction of Cx. tarsalis to feathers or controls. Responses to CO2 (5 ml/min) were low for all species (<15%) except Cx. tarsalis, which were moderate (24.5%). When feathers were combined with CO2, the resulting attraction was additive or lower than responses to feather and CO2 alone for all species except for Cx. tarsalis, which had responses that were three-fold greater than expected if responses were additive. The CO2–feather treatments were less attractive than a chicken for all species. When olfactometer assays were extended from 3 to 20 min, responses by Ae. aegypti significantly increased to a chicken and CO2 and attraction of Cx. quinquefasciatus significantly greater to chickens, CO2, and feathers. None of the volatile compounds previously identified from feathers or uropygial glands tested were attractive. Both feather-rubbed cotton balls and hexane extracts of feathers were attractive and as attractive as feathers; however, ether extracts were not attractive. Feathers clearly contribute to the attraction of host-seeking Culex spp., and future studies will focus on identification of the attractant compounds.

Aedes aegypti (Diptera: Culicidae) Biting Deterrence: Structure-Activity Relationship of Saturated and Unsaturated Fatty Acids

ABSTRACT In this study we evaluated the biting deterrent effects of a series of saturated and unsaturated fatty acids against Aedes aegypti (L), yellow fever mosquito (Diptera: Culicidae) using the K &D bioassay module system. Saturated (C6:0 to C16:0 and C18:0) and unsaturated fatty acids (C11:1 to C14:1, C16:1, C18:1, and C18:2) showed biting deterrence index (BDI) values significantly greater than ethanol, the negative control. Among the saturated fatty acids, mid chain length acids (C10:0 to C13:0) showed higher biting deterrence than short (C6:0 to C9:0) and long chain length acids (C14:0 to C18:0), except for C8:0 and C16:0 that were more active than the other short and long chain acids. The BDI values of mid chain length acids (C10:0 to C13:0) were not significantly less than N, N-diethyl-meta-toluamide (DEET), the positive control. Among the unsaturated fatty acids, C11:1 showed the highest activity (BDI = 1.05) and C18:2 had the lowest activity (BDI = 0.7). In C11:1, C12:1, and C14:1 BDI values were not significantly less than DEET. After the preliminary observations, residual activity bioassays were performed on C11:0, C12:0, C11:1, and C12:1 over a 24-h period. All the fatty acids (C11:0, C12:0, C11:1, and C12:1) and DEET showed significantly higher activity at all test intervals than the solvent control. At treatment and 1-h posttreatment, all fatty acids showed proportion not biting (PNB) values not significantly less than DEET. At 3-, 6-, and 12-h posttreatment, all fatty acids showed PNB values significantly greater than DEET. At 24-h posttreatment, only the PNB value for C12:0 was significantly higher than DEET. The dose-responses of C12:0 and DEET were determined at concentrations of 5–25 nmol/cm2. As in the residual activity bioassays, the PNB values for C12:0 and DEET at 25 nmol/cm2 were not significantly different. However, at lower concentrations, the PNB values for C12:0 were significantly greater than DEET. These results clearly indicate that mid chain length fatty acids not only have levels of biting deterrence similar to DEET at 25 nmol/cm2 in our test system, but also appeared to be more persistent than DEET. In contrast, in vivo cloth patch assay system showed that the mid-chain length fatty acids, C11:0, C11:1 C12:0, and C12:1 had minimum effective dose (MED) values greater than DEET against Ae. aegypti and their relative repellency varied according to species tested. The MED values of 120 (C11:0), 145 (C12:0) and 116 (C11:1) nmol/cm2 against Anopheles quadrimaculatus Say, indicated that these acids were not as potent as DEET with a MED of 54 nmol/cm2. The MED ratio of the C11:0 and C11:1 for all three mosquito species indicated the C11 saturated and unsaturated acids as more repellent than their corresponding C12:0 and C12:1 homologues.

Essential oils of Cupressus funebris, Juniperus communis, and J. chinensis (Cupressaceae) as repellents against ticks (Acari: Ixodidae) and mosquitoes (Diptera: Culicidae) and as toxicants against mosquitoes

ABSTRACT: Juniperus communis leaf oil, J. chinensis wood oil, and Cupressus funebris wood oil (Cupressaceae) from China were analyzed by gas chromatography and gas chromatography-mass spectrometry We identified 104 compounds, representing 66.8–95.5% of the oils. The major components were: &agr;-pinene (27.0%), &agr;-terpinene (14.0%), and linalool (10.9%) for J. communis; cuparene (11.3%) and &dgr;-cadinene (7.8%) for J. chinensis; and &agr;-cedrene (16.9%), cedrol (7.6%), and &bgr;-cedrene (5.7%) for C. funebris. The essential oils of C. funebris, J. chinensis, and J. communis were evaluated for repellency against adult yellow fever mosquitoes, Aedes aegypti (L.), host-seeking nymphs of the lone star tick, Amblyomma americanum (L.), and the blacklegged tick, Ixodes scapularis Say, and for toxicity against Ae. aegypti larvae and adults, all in laboratory bioassays. All the oils were repellent to both species of ticks. The EC95 values of C. funebris, J. communis, and J. chinensis against A. americanum were 0.426, 0.508, and 0.917 mg oil/cm2 filter paper, respectively, compared to 0.683 mg deet/cm2 filter paper. All I. scapularis nymphs were repelled by 0.103 mg oil/cm2 filter paper of C. funebris oil. At 4 h after application, 0.827 mg oil/cm2 filter paper, C. funebris and J. chinensis oils repelled ≥80% of A. americanum nymphs. The oils of C. funebris and J. chinensis did not prevent female Ae. aegypti from biting at the highest dosage tested (1.500 mg/cm2). However, the oil of J. communis had a Minimum Effective Dosage (estimate of ED99) for repellency of 0.029 ± 0.018 mg/cm2; this oil was nearly as potent as deet. The oil of J. chinensis showed a mild ability to kill Ae. aegypti larvae, at 80 and 100% at 125 and 250 ppm, respectively.

Anointing Chemicals and Hematophagous Arthropods: Responses by Ticks and Mosquitoes to Citrus (Rutaceae) Peel Exudates and Monoterpene Components

Some birds and mammals roll on or wipe themselves with the fruits or leaves of Citrus spp. or other Rutaceae. These anointing behaviors, as with anointing in general, are thought to function in the topical acquisition of chemicals that deter consumers, including hematophagous arthropods. We measured avoidance and other responses by nymphal lone star ticks (Amblyomma americanum) and adult female yellow fever mosquitoes (Aedes aegypti) to lemon peel exudate and to 24 volatile monoterpenes (racemates and isomers), including hydrocarbons, alcohols, aldehydes, acetates, ketones, and oxides, present in citrus fruits and leaves in order to examine their potential as arthropod deterrents. Ticks allowed to crawl up vertically suspended paper strips onto a chemically treated zone avoided the peel exudate and geraniol, citronellol, citral, carveol, geranyl acetate, α-terpineol, citronellyl acetate, and carvone. Ticks confined in chemically treated paper packets subsequently were impaired in climbing and other behaviors following exposure to the peel exudate and, of the compounds tested, most impaired to carveol. Mosquitoes confined in chambers with chemically treated feeding membranes landed and fed less, and flew more, when exposed to the peel exudate than to controls, and when exposed to aldehydes, oxides, or alcohols versus most hydrocarbons or controls. However, attraction by mosquitoes in an olfactometer was not inhibited by either lemon peel exudate or most of the compounds we tested. Our results support the notion that anointing by vertebrates with citrus-derived chemicals deters ticks. We suggest that some topically applied compounds are converted into more potent arthropod deterrents when oxidized on the integument of anointed animals.