Priming of inducible defenses protects Norway spruce against tree-killing bark beetles.

Plants can form an immunological memory known as defense priming, whereby exposure to a priming stimulus enables quicker or stronger response to subsequent attack by pests and pathogens. Such priming of inducible defenses provides increased protection and reduces allocation costs of defense. Defense priming has been widely studied for short-lived model plants such as Arabidopsis, but little is known about this phenomenon in long-lived plants like spruce. We compared the effects of pretreatment with sublethal fungal inoculations or application of the phytohormone methyl jasmonate (MeJA) on the resistance of 48-year-old Norway spruce (Picea abies) trees to mass attack by a tree-killing bark beetle beginning 35 days later. Bark beetles heavily infested and killed untreated trees but largely avoided fungus-inoculated trees and MeJA-treated trees. Quantification of defensive terpenes at the time of bark beetle attack showed fungal inoculation induced 91-fold higher terpene concentrations compared with untreated trees, whereas application of MeJA did not significantly increase terpenes. These results indicate that resistance in fungus-inoculated trees is a result of direct induction of defenses, whereas resistance in MeJA-treated trees is due to defense priming. This work extends our knowledge of defense priming from model plants to an ecologically important tree species.

Composition of Strawberry Floral Volatiles and their Effects on Behavior of Strawberry Blossom Weevil, Anthonomus rubi.

The strawberry blossom weevil (SBW), Anthonomus rubi, is a major pest in strawberry fields throughout Europe. Traps baited with aggregation pheromone are used for pest monitoring. However, a more effective lure is needed. For a number of pests, it has been shown that the attractiveness of a pheromone can be enhanced by host plant volatiles. The goal of this study was to explore floral volatile blends of different strawberry species (Fragaria x ananassa and Fragaria vesca) to identify compounds that might be used to improve the attractiveness of existing lures for SBW. Floral emissions of F. x a. varieties Sonata, Beltran, Korona, and of F. vesca, were collected by both solid-phase microextraction (SPME) and dynamic headspace sampling on Tenax. Analysis by gas chromatography/mass spectrometry showed the floral volatiles of F. x ananassa. and F. vesca were dominated by aromatic compounds and terpenoids, with 4-methoxybenzaldehyde (p-anisaldehyde) and α-muurolene the major compounds produced by the two species, respectively. Multi-dimensional scaling analyses separated the blends of the two species and explained differences between F. vesca genotypes and, to some degree, variation between F. x ananassa varieties In two-choice behavioral tests, SBW preferred odors of flowering strawberry plants to those of non-flowering plants, but weevils did not discriminate between odors from F. x ananassa and F. vesca flowering plants. Adding blends of six synthetic flower volatiles to non-flowering plants of both species increased the preference of SBW for these over the plants alone. When added individually to non-flowering plants, none of the components increased the preference of SBW, indicating a synergistic effect. However, SBW responded to 1,4-dimethoxybenzene, a major component of volatiles from F. viridis, previously found to synergize the attractiveness of the SBW aggregation pheromone in field studies.

Identification of (Z)-8-Heptadecene and n-Pentadecane as Electrophysiologically Active Compounds in Ophrys insectifera and Its Argogorytes Pollinator.

Sexually deceptive orchids typically depend on specific insect species for pollination, which are lured by sex pheromone mimicry. European Ophrys orchids often exploit specific species of wasps or bees with carboxylic acid derivatives. Here, we identify the specific semiochemicals present in O. insectifera, and in females of one of its pollinator species, Argogorytes fargeii. Headspace volatile samples and solvent extracts were analysed by GC-MS and semiochemicals were structurally elucidated by microderivatisation experiments and synthesis. (Z)-8-Heptadecene and n-pentadecane were confirmed as present in both O. insectifera and A. fargeii female extracts, with both compounds being found to be electrophysiologically active to pollinators. The identified semiochemicals were compared with previously identified Ophrys pollinator attractants, such as (Z)-9 and (Z)-12-C27-C29 alkenes in O. sphegodes and (Z)-9-octadecenal, octadecanal, ethyl linoleate and ethyl oleate in O. speculum, to provide further insights into the biosynthesis of semiochemicals in this genus. We propose that all these currently identified Ophrys semiochemicals can be formed biosynthetically from the same activated carboxylic acid precursors, after a sequence of elongation and decarbonylation reactions in O. sphegodes and O. speculum, while in O. insectifera, possibly by decarbonylation without preceding elongation.

The repellency and toxicity effects of essential oils from the Libyan plants Salvadora persica and Rosmarinus officinalis against nymphs of Ixodes ricinus.

Essential oils extracted from the leaves of Libyan Rosemary (Rosmarinus officinalis L.), and Miswak (Salvadora persica L.) were evaluated for their acaricidal and repellent effects on Ixodes ricinus L. nymphs (Acari: Ixodidae) using a bioassay based on an 'open filter paper method'. Rosmarinus officinalis leaf essential oil diluted to 0.5 and 1 µl/cm2 in acetone exhibited, respectively, 20 and 100% tick mortality after about 5 h of exposure. A total of 50 and 95% of I. ricinus nymphs were killed by direct contact with the oil when exposed to lethal concentrations (LC) of 0.7 µl/cm2 (LC50) and 0.95 µl/cm2 (LC95), respectively. The LC50 (0.5 µl/cm2) was reached before the end of the first 24 h of exposure time (ET), as tick mortality at 24 h was 60%. Salvadora persica leaf essential oil at 1 µl/cm2 showed a significant repellency effect against I. ricinus nymphs at 1.5 h ET. A 95% repellency was observed at a repellent concentration (RC95) of 1 µl/cm2 of S. persica, but no significant mortality was recorded at this dose of S. persica oil. Gas chromatography-mass spectrometry analyses showed that the main monoterpenes in both oils were 1,8-cineol, α-pinene, and ß-pinene, although in markedly different proportions. These results suggest that essential oils have substantial potential as alternative approaches for I. ricinus tick control.

Antifeedants Produced by Bacteria Associated with the Gut of the Pine Weevil Hylobius abietis.

The pine weevil, Hylobius abietis, is a severe forest pest insect as it feeds on newly planted conifer seedlings. To identify and develop an antifeedant could be one step towards the protection of seedlings from feeding damage by the pine weevil. With the aim to trace the origin of the antifeedants previously found in feces of the pine weevil, we investigated the culturable bacteria associated with the gut and identified the volatiles they produced. Bacterial isolates were identified by 16S ribosomal RNA gene analysis. The volatile emissions of selected bacteria, cultivated on NB media or on the grated phloem of Scots pine twigs dispersed in water, were collected and analyzed by solid-phase microextraction gas chromatography-mass spectrometry. The bacterial isolates released a variety of compounds, among others 2-methoxyphenol, 2-phenylethanol, 3-methyl-1-butanol, 1-octen-3-ol, 3-octanone, dimethyl disulfide, and dimethyl trisulfide. A strong antifeedant effect was observed by 2-phenylethanol, which could thus be a good candidate for use to protect planted conifer seedlings against feeding damage caused by H. abietis.

Sulfate turpentine: a resource of tick repellent compounds.

Compounds with tick (Ixodes ricinus) repellent properties were isolated from sulfate turpentine consisting of Norway spruce (80%) and Scots pine (20%) from southern Sweden. The turpentine was divided into two fractions by distillation under reduced pressure resulting in one monoterpene hydrocarbon fraction and a residual containing higher boiling terpenoids. The monoterpene fraction was further oxidized with SeO2 to obtain oxygenated monoterpenes with potential tick repellent properties. The oxidized fraction and the high boiling distillation residual were each separated by medium pressure liquid chromatography. The fractions were tested for tick repellency and the compounds in those with highest tick repellency were identified by GC-MS. The fractions with highest repellency contained, mainly (-)-borneol, and mixtures of (+)- and (-)-1-terpineol and terpinen-4-ol. The enantiomers of borneol showed similar tick repellent properties.

Essential oils of aromatic Egyptian plants repel nymphs of the tick Ixodes ricinus (Acari: Ixodidae).

Due to the role of Ixodes ricinus (L.) (Acari: Ixodidae) in the transmission of many serious pathogens, personal protection against bites of this tick is essential. In the present study the essential oils from 11 aromatic Egyptian plants were isolated and their repellent activity against I. ricinus nymphs was evaluated Three oils (i.e. Conyza dioscoridis L., Artemisia herba-alba Asso and Calendula officinalis L.) elicited high repellent activity in vitro of 94, 84.2 and 82%, respectively. The most active essential oil (C. dioscoridis) was applied in the field at a concentration of 6.5 µg/cm2 and elicited a significant repellent activity against I. ricinus nymphs by 61.1%. The most repellent plants C. dioscoridis, C. officinalis and A. herba-alba yielded essential oils by 0.17, 0.11 and 0.14%, respectively. These oils were further investigated using gas chromatography-mass spectrometry analysis. α-Cadinol (10.7%) and hexadecanoic acid (10.5%) were the major components of C. dioscoridis whereas in C. officinalis, α-cadinol (21.2%) and carvone (18.2%) were major components. Artemisia herba-alba contained piperitone (26.5%), ethyl cinnamate (9.5%), camphor (7.7%) and hexadecanoic acid (6.9%). Essential oils of these three plants have a potential to be used for personal protection against tick bites.

The gut microbiota of the pine weevil is similar across Europe and resembles that of other conifer-feeding beetles.

The pine weevil (Hylobius abietis, Coleoptera: Curculionidae) is an important pest of conifer seedlings in Europe. Despite its economic importance, little is known about the composition of its gut microbial community and the role it plays in mediating the weevil's ability to utilize conifers as a food source. Here, we characterized the gut bacterial communities of different populations of H. abietis across Europe and compared them to those of other beetles that occupy similar ecological niches. We demonstrate that the microbial community of H. abietis is similar at higher taxonomic levels (family and genus) across locations in Europe, with Wolbachia as the dominant microbe, followed by Enterobacteria and Firmicutes. Despite this similarity, we observed consistent differences between countries and locations, but not sexes. Our meta-analysis demonstrates that the gut bacterial community of the pine weevil is very similar to that of bark beetles that also exploit conifers as a food source. The Enterobacteriaceae symbionts of both host taxa are especially closely related phylogenetically. Conversely, the microbiota of H. abietis is distinct from that of closely related weevils feeding on nonconifer food sources, suggesting that the microbial community of the pine weevil is determined by the environment and may be relevant to host ecology. Furthermore, several H. abietis-associated members of the Enterobacteriaceae family are known to contain genes involved in terpenoid degradation. As such, we hypothesize that the gut microbial community is important for the utilization of conifer seedlings as a food source, either through the detoxification of plant secondary metabolites or through the supplementation of essential nutrients.

Gravid Anopheles gambiae sensu stricto avoid ovipositing in Bermuda grass hay infusion and it's volatiles in two choice egg-count bioassays.

BACKGROUND: A number of mosquito species in the Culex and Aedes genera prefer to lay eggs in Bermuda grass (Cynodon dactylon) hay infusions compared to water alone. These mosquitoes are attracted to volatile compounds from the hay infusions making the infusions effective baits in gravid traps used for monitoring vectors of arboviral and filarial pathogens. Since Bermuda grass is abundant and widespread, it is plausible to explore infusions made from it as a potential low cost bait for outdoor monitoring of the elusive malaria vector Anopheles gambiae s.s. METHODS: This study investigated preferential egg laying of individual An. gambiae s.s. in hay infusion or in tap water treated with volatiles detected in hay infusion headspace compared to tap water alone, using two-choice egg-count bioassays. Infusions were prepared by mixing 90 g of dried Bermuda grass (hay) with 24 L of unchlorinated tap water in a bucket, and leaving it for 3 days at ambient temperature and humidity. The volatiles in the headspace of the hay infusion were sampled with Tenax TA traps for 20 h and analysed using gas chromatography coupled to mass spectrometry. RESULTS: In total, 18 volatiles were detected in the infusion headspace. Nine of the detected compounds and nonanal were selected for bioassays. Eight of the selected compounds have previously been suggested to attract/stimulate egg laying in An. gambiae s.s. Gravid females were significantly (p < 0.05) less likely to lay eggs in hay infusion dilutions of 25, 50 and 100 % and in tap water containing any of six compounds (3-methylbutanol, phenol, 4-methylphenol, nonanal, indole, and 3-methylindole) compared to tap water alone. The oviposition response to 10 % hay infusion or any one of the remaining four volatiles (4-hepten-1-ol, phenylmethanol, 2-phenylethanol, or 4-ethylphenol) did not differ from that in tap water. CONCLUSIONS: Anopheles gambiae s.s. prefers to lay eggs in tap water rather than Bermuda grass hay infusion. This avoidance of the hay infusion appears to be mediated by volatile organic compounds from the infusion. It is, therefore, unlikely that Bermuda grass hay infusion as formulated and used in gravid traps for Culex and Aedes mosquitoes will be suitable baits for monitoring gravid An. gambiae s.s.

Cedrol, a malaria mosquito oviposition attractant is produced by fungi isolated from rhizomes of the grass Cyperus rotundus.

BACKGROUND: Cedrol, a sesquiterpene alcohol, is the first identified oviposition attractant for African malaria vectors. Finding the natural source of this compound might help to elucidate why Anopheles gambiae and Anopheles arabiensis prefer to lay eggs in habitats containing it. Previous studies suggest that cedrol may be a fungal metabolite and the essential oil of grass rhizomes have been described to contain a high amount of different sesquiterpenes. RESULTS: Rhizomes of the grass Cyperus rotundus were collected in a natural malaria mosquito breeding site. Two fungi were isolated from an aqueous infusion with these rhizomes. They were identified as Fusarium falciforme and a species in the Fusarium fujikuroi species complex. Volatile compounds were collected from the headspace above fungal cultures on Tenax traps which were analysed by gas chromatography-mass spectrometry (GCMS). Cedrol and a cedrol isomer were detected in the headspace above the F. fujikuroi culture, while only cedrol was detected above the F. falciforme culture. CONCLUSION: Cedrol an oviposition attractant for African malaria vectors is produced by two fungi species isolated from grass rhizomes collected from a natural mosquito breeding site.

Methyl Jasmonate-Induced Monoterpenes in Scots Pine and Norway Spruce Tissues Affect Pine Weevil Orientation.

In large parts of Europe, insecticide-free measures for protecting conifer plants are desired to suppress damage by the pine weevil Hylobius abietis (L.). Treatment with methyl jasmonate (MeJA), a chemical elicitor already used in crop production, may enhance expression of chemical defenses in seedlings in conifer regenerations. However, in a previous experiment, MeJA treatment resulted in substantially better field protection for Scots pine (Pinus sylvestris L.) than for Norway spruce (Picea abies (L.) Karst.). Hypothesizing that the variations may be at least due partly to volatiles released by MeJA-treated seedlings and their effects on pine weevil orientation, we examined tissue extracts of seedlings (from the same batches as previously used) by two-dimensional GC-MS. We found that the MeJA treatment increased contents of the monoterpene (-)-ß-pinene in phloem (the weevil's main target tissue) of both tree species, however, the (-)-ß-pinene/(-)-α-pinene ratio increased more in the phloem of P. sylvestris. We also tested the attractiveness of individual monoterpenes found in conifer tissues (needles and phloem) for pine weevils using an arena with traps baited with single-substance dispensers and pine twigs. Trap catches were reduced when the pine material was combined with a dispenser releasing (-)-ß-pinene, (+)-3-carene, (-)-bornyl acetate or 1,8-cineole. However, (-)-α-pinene did not have this effect. Thus, the greater field protection of MeJA-treated P. sylvestris seedlings may be due to the selective induction of increases in contents of the deterrent (-)-ß-pinene, in contrast to strong increases in both non-deterrent (-)-α-pinene and the deterrent (-)-ß-pinene in P. abies seedlings.

Discovery of an oviposition attractant for gravid malaria vectors of the Anopheles gambiae species complex.

BACKGROUND: New strategies are needed to manage malaria vector populations that resist insecticides and bite outdoors. This study describes a breakthrough in developing 'attract and kill' strategies targeting gravid females by identifying and evaluating an oviposition attractant for Anopheles gambiae s.l. METHODS: Previously, the authors found that gravid An. gambiae s.s. females were two times more likely to lay eggs in lake water infused for six days with soil from a natural oviposition site in western Kenya compared to lake water alone or to the same but autoclaved infusion. Here, the volatile chemicals released from these substrates were analysed with a gas-chromatograph coupled to a mass-spectrometer (GC-MS). Furthermore, the behavioural responses of gravid females to one of the compounds identified were evaluated in dual choice egg-count bioassays, in dual-choice semi-field experiments with odour-baited traps and in field bioassays. RESULTS: One of the soil infusion volatiles was readily identified as the sesquiterpene alcohol cedrol. Its widespread presence in natural aquatic habitats in the study area was confirmed by analysing the chemical headspace of 116 water samples collected from different aquatic sites in the field and was therefore selected for evaluation in oviposition bioassays. Twice as many gravid females were attracted to cedrol-treated water than to water alone in two choice cage bioassays (odds ratio (OR) 1.84; 95% confidence interval (CI) 1.16-2.91) and in experiments conducted in large-screened cages with free-flying mosquitoes (OR 1.92; 95% CI 1.63-2.27). When tested in the field, wild malaria vector females were three times more likely to be collected in the traps baited with cedrol than in the traps containing water alone (OR 3.3; 95% CI 1.4-7.9). CONCLUSION: Cedrol is the first compound confirmed as an oviposition attractant for gravid An. gambiae s.l. This finding paves the way for developing new 'attract and kill strategies' for malaria vector control.

Fungal Symbionts of the Spruce Bark Beetle Synthesize the Beetle Aggregation Pheromone 2-Methyl-3-buten-2-ol.

Tree-killing bark beetles depend on aggregation pheromones to mass-attack their host trees and overwhelm their resistance. The beetles are always associated with phytopathogenic ophiostomatoid fungi that probably assist in breaking down tree resistance, but little is known about if or how much these fungal symbionts contribute to the beetles' aggregation behavior. In this study, we determined the ability of four major fungal symbionts of the spruce bark beetle Ips typographus to produce beetle aggregation pheromones. The fungi were incubated on Norway spruce Picea abies bark, malt agar, or malt agar amended with 0.5% (13)C glucose. Volatiles present in the headspace of each fungus were analyzed for 7 days after incubation using a SPME autosampler coupled to a GC/MS. Two Grosmannia species (G. penicillata and G. europhioides) produced large amounts of 2-methyl-3-buten-2-ol (MB), the major component in the beetles' aggregation pheromone blend, when growing on spruce bark or malt agar. Grosmannia europhioides also incorporated (13)C glucose into MB, demonstrating that the fungi can synthesize MB de novo using glucose as a carbon source. This is the first clear evidence that fungal symbionts of bark beetles can produce components in the aggregation pheromone blend of their beetle vectors. This provides new insight into the possible ecological roles of fungal symbionts in bark beetle systems and may deepen our understanding of species interactions and coevolution in these important biological systems.

Penicillium expansum volatiles reduce pine weevil attraction to host plants.

The pine weevil Hylobius abietis (L.) is a severe pest of conifer seedlings in reforested areas of Europe and Asia. To identify minimally toxic and ecologically sustainable compounds for protecting newly planted seedlings, we evaluated the volatile metabolites produced by microbes isolated from H. abietis feces and frass. Female weevils deposit feces and chew bark at oviposition sites, presumably thus protecting eggs from feeding conspecifics. We hypothesize that microbes present in feces/frass are responsible for producing compounds that deter weevils. Here, we describe the isolation of a fungus from feces and frass of H. abietis and the biological activity of its volatile metabolites. The fungus was identified by morphological and molecular methods as Penicillium expansum Link ex. Thom. It was cultured on sterilized H. abietis frass medium in glass flasks, and volatiles were collected by SPME and analyzed by GC-MS. The major volatiles of the fungus were styrene and 3-methylanisole. The nutrient conditions for maximum production of styrene and 3-methylanisole were examined. Large quantities of styrene were produced when the fungus was cultured on grated pine bark with yeast extract. In a multi-choice arena test, styrene significantly reduced male and female pine weevils' attraction to cut pieces of Scots pine twigs, whereas 3-methylanisole only reduced male weevil attraction to pine twigs. These studies suggest that metabolites produced by microbes may be useful as compounds for controlling insects, and could serve as sustainable alternatives to synthetic insecticides.

Antitermite activity of beta-caryophyllene epoxide and episulfide.

Caryophyllene-6,7-epoxide and caryophyllene-6,7-episulfide can be easily synthesized from beta-caryophyllene by autoxidation or episulfidation. The bioactivities of beta-caryophyllene and its derivatives were investigated against the subterranean termite Reticulitermes speratus Kolbe. The antifeedant, feeding, and termiticidal activities of each compound were tested using no-choice, dual-choice, and non-contact methods. Antitermitic activities were not shown by beta-caryophyllene, but were observed for the oxide and sulfide derivatives. Caryophyllene-6,7-episulfide showed especially high antifeedant and termiticidal activities. Thus, naturally abundant, non-bioactive beta-caryophyllene can be easily converted into an antitermite reagent via a non-biological process.

UV-B exposure of indoor-grown Picea abies seedlings causes an epigenetic effect and selective emission of terpenes.

Terpenoids are involved in various defensive functions in plants, especially conifers. Epigenetic mechanisms, for example DNA methylation, can influence plant defence systems. The purpose of the present study was to investigate the influence of UV-B exposure on the release of terpenoids from spruce seedlings and on needle DNA methylation. Ten-week-old seedlings grown indoors were exposed to UV-B radiation during 4 h, and the volatile compounds emitted from the seedlings were analysed. Analysis of the volatiles 1, 3, and 22 d after this UV-B exposure showed that bornyl acetate, borneol, myrcene, and limonene contents increased during the first 3 days, while at day 22 the level of emission had returned to the control level. UV-B exposure decreased the level of DNA methylation in needles of young seedlings, reflected in methylation changes in CCGG sequences. Exposure of young seedlings to UV-B radiation might be a way to potentiate the general defensive capacity, improving their ability to survive in outdoor conditions. UV-B-induced defence is discussed in the light of epigenetic mechanisms.

p-Cresol: a sex pheromone component identified from the estrous urine of mares.

Previously it was shown that m- and p-cresols in the urine of mares exhibits a temporally reproducible pattern that is dependent on ovarian activity and, thus, provides information about the timing of ovulation. New behavioral data demonstrate 1) that stallions spend significantly more time sniffing p-cresol as compared to o-, and m-cresols, and, 2) that the extent of stallions' erections differ significantly in response to different types of samples. The lowest erection level was recorded for the pure-water control, a moderate erection level was elicited by the urine of diestrous mares, and the highest erection level was elicited by urine of a diestrous mare containing synthetic p-cresol at a quantity equivalent to half of the amount of p-cresol found in the urine of estrous mares. Consequently, p-cresol is at least one of the components of a horse sex pheromone.

Timing of male sex pheromone biosynthesis in a butterfly - different dynamics under direct or diapause development.

The life history traits and behavior of the butterfly Pieris napi are well-known, as the species is often used as a model organism for evolutionary and ecological studies. The species has two or more generations per year in the major part of its temperate distribution, and as different selection pressures affect the different generations, both behavioral and physiological seasonal polyphenisms have been shown previously. Here, we explored the dynamics of male sex pheromone production. The two generations are shown to have significantly different scent compositions early in life; the direct developers--who have shorter time for pupal development--need the first 24 hr of adult life after eclosion to synthesize the sex pheromone citral (geranial and neral 1:1)--whereas the diapausing individuals who have spent several months in the pupal stage eclose with adult scent composition. Resource allocation and biosynthesis also were studied in greater detail by feeding butterflies (13)C labeled glucose either in the larval or adult stage, and recording incorporation into geranial, neral, and other volatiles produced. Results demonstrate that the pheromone synthesized by newly eclosed adult males is based on materials ingested in the larval stage, and that adult butterflies are able to synthesize the pheromone components geranial and neral and the related alcohols also from adult intake of glucose. In summary, our study shows that time-stress changes the timing in biosynthesis of the complete pheromone between generations, and underpins the importance of understanding resource allocation and the physiological basis of life history traits.

Mosquito (Diptera: Culicidae) repellency field tests of essential oils from plants traditionally used in Laos.

Essential oils of Hyptis suaveolens (Lamiaceae), Croton roxburghii (Euphorbiaceae), and Litsea cubeba (Lauraceae) were tested in the field near Vientiane city, Lao PDR, on humans for repellent activity against mosquitoes. Landing mosquitoes were collected and later identified. The most abundant mosquitoes captured belonged to the genera Armigeres, Culex, and Aedes. All the plant oils tested at concentrations of 1.7 microg/cm(2), 3.3 microg/cm(2), and 6.3 microg/cm(2) were significantly more mosquito repellent than the negative control. Croton oil was significantly repellent against mosquitoes of the three genera at the highest (6.3 microg/cm(2)) concentration tested. Litsea oil was significantly repellent against Armigeres at all (1.7 microg/cm(2), 3.3 microg/cm(2), and 6.3 microg/cm(2)) concentrations tested. Hyptis oil was significantly repellent against Armigeres at 3.3 microg/cm(2) and 6.3 microg/cm(2) and against Culex at 1.7 microg/cm(2) and 6.3 microg/cm(2). The oils were analyzed for chemical content of volatiles, mainly terpenes. Main constituents were beta-pinene, sabinene, and 1,8-cineol from oils of the green parts of H. suaveolens; alpha-pinene, beta-pinene, and alpha-phellandrene from fresh bark of C. roxburghii; and alpha-pinene, beta-phellandrene, sabinene, and 1,8-cineol from fresh fruits of L. cubeba.

Chemical composition and repellency of essential oils from four medicinal plants against Ixodes ricinus nymphs (Acari: Ixodidae).

In our search for effective tick repellents from plant origin, we investigated the effect of essential oils of four medicinal and culinary plants belonging to the family Lamiaceae on nymphs of the tick Ixodes ricinus (L.). The essential oils of the dry leaves of Rosmarinus officinalis (Rosemary) (L.), Mentha spicata (Spearmint) (L.), Origanum majorana (Majoram) (L.), and Ocimum basilicum (Basil) (L.) were isolated by steam distillation and 15 microg/cm2 concentration of oils was tested against ticks in a laboratory bioassay. The oils of R. officinalis, M. spicata, and O. majorana showed strong repellency against the ticks 100, 93.2, and 84.3%, respectively, whereas O. basilicum only showed 64.5% repellency. When tested in the field, the oils of R. officinalis and M. spicata showed 68.3 and 59.4% repellency at a concentration of 6.5 microg/cm2 on the test cloths. The oils were analyzed by gas chromatography mass spectrometry and the major compounds from the most repellent oils were 1,8-cineole, camphor, linalool, 4-terpineol, borneol, and carvone.