Mimicry of queen Dufour's gland secretions by workers of Apis mellifera scutellata and A. m. capensis

The development of the Dufour's gland of workers of the two honey bee races Apis mellifera scutellata and A. m. capensis was measured. The Dufour's glands of A. m. capensis workers were longer and increased in length more rapidly than the glands of workers of A. m. scutellata at comparable ages. Analysis of the Dufour's gland secretions of workers and queens of both races revealed that there were caste and racial differences. Secretions of queenright A. m. scutellata workers were dominated by a series of long-chain hydrocarbons. In contrast the secretions of the A. m. capensis workers both under queenright and queenless conditions were a mixture of hydrocarbons and wax-type esters, as were those of queens. Multivariate analysis of the secretion profiles indicated that laying workers of both races mimic queens. The secretions of the A. m. capensis laying workers mimicked queen secretions most closely, enabling them to act as successful social parasites.     

Nestmate recognition in social wasps: manipulation of hydrocarbon profiles induces aggression in the European hornet

The influence of individual cuticular hydrocarbons on nestmate recognition in the European hornet, Vespa crabro L., was investigated. We observed the behavioural response of workers towards differently treated dead conspecifics in a bioassay. Dummies were extracted with dichloromethane and extracts were spiked with microgram amounts of synthetic hydrocarbons naturally occurring on the cuticle of V. crabro. These modified extracts were reapplied to extracted workers that were subsequently tested in the bioassay. Non-spiked nestmate dummies (negative control) and untreated non-nestmate dummies (positive control) were tested in control experiments. The addition of only heneicosane or a mixture of heneicosane, tricosane, and (Z)-9-tricosene to the extracts led to a significant increase of agonistic behaviour in workers leaving the nest for foraging flights. Returning workers reacted much less aggressively than those leaving. This is one of the first behavioural proofs that manipulation of cuticular hydrocarbon profiles can be perceived by a social insect species. The results support the hypothesis that colony-specific cuticular hydrocarbon profiles are involved in the phenomenon of nestmate recognition among social insects.     

Colony disassociation following diet partitioning in a unicolonial ant

Discriminating nestmates from alien conspecifics via chemical cues is recognized as a critical element in maintaining the integrity of insect societies. We determined, in laboratory experiments, that nestmate recognition in an introduced population of the Argentine ant, Linepithema humile, is modified by hydrocarbons acquired from insect prey, and that workers from spatially isolated colony fragments, each provided with prey that possessed distinct cuticular hydrocarbons, displayed aggressive behavior towards their former nestmates. Isolation for 28 days or more between colony fragments fed different prey was sufficient to prevent re-establishment of inter-nest communication for at least an additional 28 days through the introduction of a bridge between the nests. Ants possessed intrinsic cuticular hydrocarbons plus only those hydrocarbons from the prey they received during the isolation period. Colony fragments which were isolated for less than 28 days reunited with workers possessing both prey hydrocarbons. Therefore, L. humile nestmate recognition may be dynamic, being in part dependent on the spatio-temporal distribution of prey, along with physical factors permitting or restricting access of subcolony units to those prey.     

Postmating changes in cuticular chemistry and visual appearance in <Emphasis Type="Italic">Ectatomma tuberculatum</Emphasis> queens (Formicidae: Ectatomminae)

In the ectatommine ant Ectatomma tuberculatum, the visual appearance of queens changes after mating and ovarian development in that their cuticle turns from shiny to matte. In this study, we have shown that this change seems to be caused by 15-fold accumulation of hydrocarbons, in particular heptacosane that covers the multiple grooves present on the cuticular surface creating a wax coat in mated fully fertile queens. Analyses of the scrapped wax revealed that it is composed largely of heptacosane. Peak-by-peak comparison of the cuticular hydrocarbon (CHC) composition of mated, virgin with developed ovaries and virgin with nondeveloped ovaries revealed significant differences between the queen groups. Although the total amount of the CHC of virgin queens with developed ovaries was not higher than virgin queens that did not have developed ovaries, the composition showed a shift toward the mated queen. While it is possible that the large accumulation of hydrocarbons may give extra physical and chemical protection to queens, we propose that the switch in the relative abundance of heptacosane and nonacosane and perhaps of other components is indicative of being a mating and fertility cue. This is the first report in social insects where external chemical changes are accompanied by changes in visual appearance.     

Surface lipids of queen-laid eggs do not regulate queen production in a fission-performing ant

In animal societies, most collective and individual decision making depends on the presence of reproductive individuals. The efficient transmission of information among reproductive and non-reproductive individuals is therefore a determinant of colony organization. In social insects, the presence of a queen modulates multiple colonial activities. In many species, it negatively affects worker reproduction and the development of diploid larvae into future queens. The queen mostly signals her presence through pheromone emission, but the means by which these chemicals are distributed in the colony are still unclear. In several ant species, queen-laid eggs are the vehicle of the queen signal. The aim of this study was to investigate whether queen-laid eggs of the ant Aphaenogaster senilis possess queen-specific cuticular hydrocarbons and/or Dufour or poison gland compounds, and whether the presence of eggs inhibited larval development into queens. Our results show that the queen- and worker-laid eggs shared cuticular and Dufour hydrocarbons with the adults; however, their poison gland compounds were not similar. Queen-laid eggs had more dimethylalkanes and possessed a queen-specific mixture of cuticular hydrocarbons composed of 3,11?+?3,9?+?3,7-dimethylnonacosane, in higher proportions than did worker-laid eggs. Even though the queen-laid eggs were biochemically similar to the queen, their addition to experimentally queenless groups did not prevent the development of new queens. More studies are needed on the means by which queen ant pheromones are transmitted in the colony, and how these mechanisms correlates with life history traits.     

Attraction of the tropical bont tick,Amblyomma variegatum,to human breath and to the breath components acetone,NO and CO2

Ticks are of medical and veterinary importance and employ several cues in search of a host. Olfaction is one modality by which ticks locate a blood-meal and breath is the major vent of gaseous and volatile metabolites from the host that could contribute to this search. We studied the responses of a hunter tick, Amblyomma variegatum, to diluted human breath and five of its components (acetone, CO₂, NO, isoprene and NH₃) while walking in an air stream on a locomotion compensator. Diluted breath elicited the greatest responses of all treatments in terms of time to onset of upwind walk, attraction, speed and local search behaviour after stimulus off. Acetone, NO and CO₂ also attracted, but with a reduced speed in the case of acetone and NO. Neither isoprene nor NH₃ induced any response. Our study indicates that breath was the most adequate stimulus tested. It also attracted two other ixodid tick species, Rhipicephalus sanguineus and Ixodes ricinus, as well as the argasid tick, Ornithodorus moubata. It appears that the evolution of resource tracking in ticks included sensory and behavioural adaptations for recognition and orientation to host metabolites that are regularly expelled in breath.     

Is the contribution of bacteria to terrestrial carbon budget greatly underestimated?

Some commonly found species of soil bacteria use low molecular weight organic acids as their sole source of carbon and energy. This study shows that acids such as citrate and oxalate (produced in large amounts by fungi and plants) can rapidly be consumed by these bacteria. Two strains, Ralstonia eutropha and Xanthobacter autotrophicus, were cultured on acetate- and citrate-rich media. The resulting CO₂ and/or HCO₃^- reacted with calcium ions to precipitate two polymorphs of calcium carbonate (CaCO₃), calcite and vaterite, depending on the quantity of slime produced by the strains. This production of primary calcium carbonate crystals by oxalate- and citrate-degrading bacteria from soil organic carbon sources highlights the existence of an important and underestimated potential carbon sink.     

The flavo-enzyme xanthine oxidase is under circadian control in the marine alga Gonyaulax

The activity of xanthine oxidoreductases (xanthine oxidase, XO, EC 1.2.3.2 and xanthine dehydrogenase, XDH, EC 1.1.1.204) in partially purified extracts of Gonyaulax polyedra was measured over 24 h both in a light:dark cycle and in constant light. This is the first demonstration of xanthine oxidoreductase in a unicellular alga. The activity of the O₂-dependent form (XO) was found to be 15 times higher in light than in darkness. The same time-of-day specific differences persisted in constant light, demonstrating a control of XO by the circadian clock. In contrast, the activity of the NAD-dependent form (XDH) is not under circadian control. Because pharmacological inhibition of XO also blocks the effect of blue light on the Gonyaulax circadian clock, the possible relationship between XO and light reception in this unicellular alga will be discussed.     

Daughters inherit colonies from mothers in the 'living-fossil' ant Nothomyrmecia macrops

Newly mated queens of monogynous (single queen) ants usually found their colonies independently, without the assistance of workers. In polygynous (multiple queen) species queens are often adopted back into their natal nest and new colonies are established by budding. We report that the Australian 'living-fossil' ant, Nothomyrmecia macrops, is exceptional in that its single queen can be replaced by one of the colony's daughters. This type of colony founding is an interesting alternative reproductive strategy in monogynous ants, which maximizes fitness under kin selection. Successive queen replacement results in a series of reproductives over time (serial polygyny), making these colonies potentially immortal. Workers raise nieces and nephews (relatedness h 0.375) the year after queen replacement. Although N. macrops is 'primitive' in many other respects, colony inheritance is likely to be a derived specialization resulting from ecological constraints on solitary founding.     

Synthetic attractants for the bark beetle parasitoid Coeloides bostrichorum Giraud (Hymenoptera: Braconidae)

Coeloides bostrichorum Giraud parasitoids (Hymenoptera: Braconidae) attack late larval stages of various bark beetle species breeding in spruce. Volatile compounds collected from Norway spruce (Picea abies) infested by Ips typographus L. (Coleoptera: Scolytidae) were analysed by coupled gas chromatography-mass spectrometry (GC-MS) and GC-electroantennographic detection (GC-EAD). Monoterpene hydrocarbons are the predominant volatile compounds of fresh Norway spruce, while the presence of oxygenated monoterpenes indicates damaged trees. Between one and eight EAD-active oxygenated monoterpenes were used, in amounts reflecting their natural abundance in spruce trees containing bark beetle larvae, to prepare five synthetic baits which were tested in wind tunnel bioassays. Odour samples collected from spruce logs containing the preferred host stage were attractive, while similar samples from uninfested logs failed to elicit any flight activity. However, when a four- or an eight-component synthetic bait was added to volatiles collected from uninfested spruce logs, this combination was as attractive as volatiles collected from infested spruce logs.     

“You are what you eat”: Diet modifies cuticular hydrocarbons and nestmate recognition in the Argentine ant, Linepithema humile

 Nestmate recognition plays a key role in the behavior and evolution of social insects. We demonstrated that hydrocarbons are the chemical cues used in Argentine ant, Linepithema humile, nestmate recognition, and that these hydrocarbons can be acquired from insect prey. Consequently, Argentine ant cuticular hydrocarbon patterns reveal the same hydrocarbons present in their diet. Diet alters both the recognition cues present on the cuticular surface and the response of nestmates to this new colony odor, resulting in aggression between former nestmates reared on different insect prey. Received: 16 June 2000 / Accepted: 14 August 2000     

Indirect interactions between a phytopathogenic and an entomopathogenic fungus

Slow growth in herbivores may lead to higher mortality, due to prolonged exposure to natural enemies. A number of studies has tested the 'slow-growth, high-mortality' hypothesis using predators or parasitoids as natural enemies of the herbivore, but the possible role of pathogens is poorly documented. We provide evidence that phytopathogenic infection of a plant enhances the susceptibility of herbivore larvae to a generalist entomopathogen. Larvae of the mustard leaf beetle Phaedon cochleariae grow more slowly when feeding on Chinese cabbage leaves infected by the phytopathogenic fungus Alternaria brassicae than larvae feeding on healthy leaves. Treatment of such larvae feeding on diseased plants with an LD₅₀ of the entomopathogenic fungus Metarhizium anisopliae resulted in 100% mortality, compared with a mortality rate of 54% in the control larvae feeding on uninfected leaves. This is the first demonstration of an interaction between a phyto- and an entomopathogenic fungus.     

Population risk assessment of ambient benzene and evaluation of benzene regulation in gasoline in Japan

The aggregate population cancer risk due to ambient benzene was evaluated for the entire Japanese population. This was done using data of ambient NO_x levels measured at as many as 2000 air pollution monitoring stations nationwide and the regression equation between the levels of benzene and NO_x obtained by continuous monitoring. The population-weighted exposure levels for the roadside population and general-environment population were calculated to be 2.1 ppb and 1.0 ppb, respectively. Eighty-four percent of the entire population was exposed to a lifetime cancer risk level of 1᎒^-5 or greater, and the annual number of cancer deaths was estimated to be 29.6 cases at the 1997 benzene level. Due to the regulation which established the upper limit of benzene content in gasoline to be 1 vol. % by January 2000, the total emission of benzene was estimated to be reduced by 27%, compared to 1993. The annual number of cancer deaths was expected to be reduced by 8.8 cases through the regulation of benzene in gasoline. Japanese petroleum companies have estimated the annual cost of the reduction of the benzene content in gasoline to be 25.6 billion yen. The cost per life saved by the regulation of benzene in gasoline is estimated to be 2.9 billion yen. The method implemented in this study to predict population risk is useful for identifying potential risk management options and evaluating their effectiveness before making decisions.     

Differential anti-lipid peroxidative activity of melatonin

Scavenging activities of melatonin, which is a pineal secretory product and functions in circadian biology, and its related compounds against reactive oxygen species such as superoxide anion radical, hydrogen peroxide, hydroxyl radical and singlet oxygen as well as organic peroxide radical (t-BuOO”) were evaluated chemically by using electron spin resonance-spin trap and chemiluminescence methods. Antioxidative activity of the compounds was estimated by IC₅₀ value (µM), 50% inhibiting concentration of a compound against reactive oxygen species formed in each system, and the second-order rate constants (k₂) for the reactions of the compounds and superoxide anion radical or hydroxyl radical. Because melatonin has exhibited the highest scavenging activity against t-BuOO”, the biochemical anti-lipid peroxide radical scavenging activities of melatonin were examined. We found that melatonin exhibits higher anti-lipid peroxidative activity in the rat brain microsomes than in the rat liver microsomal and liposomal systems, suggesting that melatonin may function as a treatment for reactive oxygen species-related diseases of the brain.     

A queen pheromone induces workers to kill sexual larvae in colonies of the red imported fire ant (<Emphasis Type="Italic">Solenopsis invicta</Emphasis>)

We conducted five bioassays to study how queens control the execution of sexual larvae by workers in colonies of the red imported fire ant, Solenopsis invicta. In each assay, subset colonies were made from many large polygyne colonies, and the 20 sexual larvae they contained were monitored over time. Sexual larvae mostly survived in queenless colonies, but were mostly killed in colonies with a single dealated queen, regardless of whether or not the queen was fertilized. The larvae were also killed when fresh corpses of queens were added to queenless colonies. Whereas acetone extracts of queens did not produce a significant increase in killings, extracts in buffered saline induced workers to execute most sexual larvae, indicating successful extraction of an execution pheromone. We identified the probable storage location of the chemical as the poison sac, and found both fresh (1 day) and old (21 day) extracts of poison sacs to be equally effective in inducing executions. The pheromone is stable at room temperature, perhaps because venom alkaloids also present in the extracts keep the pheromone from degrading. It is apparently either proteinaceous or associated with a proteinaceous molecule, a novel finding, as no queen pheromone of a proteinaceous nature has been previously demonstrated in ants.     

Scent of a queen—cuticular hydrocarbons specific for female reproductives in lower termites

In social insects, it is assumed that signals of the queen inform nestmates about her reproductive status. Thus, workers forego their own reproduction if the queen signals high fertility. In hemimetabolous termites, little is known about reproductive inhibition, but evidence exists for a royal-pair control. Workers of lower termites exhibit a high developmental flexibility and are potentially able to become reproductives, but the presence of a fertile reproductive restrains them from reaching sexual maturity. The nature of this control, however, remains unknown. Here, we report on qualitative differences in cuticular hydrocarbon profiles between queens and workers of the basal drywood termite Cryptotermes secundus. Queens were characterized by a shift to long-chained and branched hydrocarbons. Most remarkably, similar chemical patterns are regarded as fertility cues of reproductives in social Hymenoptera. This might suggest that both groups of social insects convergently evolved similar chemical signatures. The present study provides deeper insights into how termites might have socially exploited these signatures from sexual communication in their cockroach-like ancestor. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Photodynamic action of some sensitizers by photooxidation of luminol

We report the development of a novel simple experimental method which allows the comparison of new photosensitizers based on their production of reactive oxygen species. A high-performance liquid chromatography (HPLC) assay permits the monitoring of several substances (sensitizer, reactant and oxidized end product) simultaneously on a single chromatogram. Photoreactions were monitored throughout their course by the HPLC assay surveying the sensitizers' efficiency of singlet oxygen production by the oxidative decomposition of luminol. Several photosensitizers were tested: Rose Bengal, Methylene Blue, Protoporphyrin IX, Photosan III, Photofrin, Hypericin and Pseudohypericin. Additionally, photoreactions were monitored by a standard pO₂ detection system. The measurements of the two detection methods were strongly correlated. Rose Bengal proved to be the most efficient photosensitizer, clearly decreasing the luminol concentration and causing a corresponding increase in aminophthalic acid. Our experiments show that when factors necessary for photochemical reactions are absent or are blocked (antioxidants), no reaction can be detected.     

Hydrocarbon synthesis by enzymatically dissociated oenocytes of the abdominal integument of the German Cockroach,<Emphasis Type="Italic"> Blattella germanica</Emphasis>

In insects, hydrocarbons waterproof the cuticle, protect the insect from the external environment, and serve as semiochemicals or their metabolic precursors. In the German cockroach, Blattella germanica, hydrocarbons are synthesized by the abdominal integument, but the precise site of biosynthesis is not known. We developed a method for separation of oenocytes from other cells in the abdominal integument using enzymatic dissociation followed by Percoll gradient centrifugation. Radiolabeled propionate was then used to monitor de novo synthesis of hydrocarbons by dissociated cells. Oenocyte-enriched cell suspensions of abdominal sternites synthesized hydrocarbons, whereas suspensions enriched with epidermal cells did not. Our results show conclusively that hydrocarbons are produced by oenocytes not only in insects whose oenocytes are localized within the hemocoel, but also in those insects whose oenocytes are within the abdominal integument. Furthermore, these data support a hemolymph pathway for transport and delivery of hydrocarbons to both external and internal tissues, including the epicuticle, fat body, and ovaries.     

Inverse correlation between mitochondrial size and metabolic competence: a quantitative cytochemical study of cytochrome oxidase activity

Mitochondria are topologically closed bilayered systems where the synthesis of adenosine triphosphate (ATP) from adenosine diphosphate (ADP) and inorganic phosphate occurs via oxidative phosphorylation. The ordered architecture (and its extension) of the mitochondria (i.e. inner membrane, outer membrane and cristae) constitutes a critical topographic arrangement for their energy-providing mechanisms. Thus, quantitative estimations of the ultrastructural features of organelles preferentially stained by means of function-related cytochemical reactions reliably report on their potential to supply adequate amounts of ATP. On the basis of this rationale, we carried out a computer-assisted cytochemical study of cytochrome oxidase (COX) activity on mitochondria of different size in the cerebellar cortex of adult rats. The total intra-mitochondrial area of the cytochemical precipitates (CPA)/mitochondrion, the area (MA) and the longer diameter (F_max) of COX-positive organelles were measured. The ratio (R): CPA/MA was also calculated and referred to as the percentage of mitochondrial inner membrane area involved in COX activity. The regression analysis of R vs MA showed a significant inverse correlation (r=т.905). The fourfold increase in MA from quartiles I to IV was matched by increases in F_max and CPA, respectively, but it was also related to a 25% decrease in R. By matching quantitative cytochemical estimations of COX activity within mitochondria with the morphometric assessment of their ultrastructural features, the present study correlates size to the metabolic competence of COX-positive organelles. Quantitative cytochemistry of COX activity is currently regarded as a reliable marker of cellular metabolism; thus our findings support the hypothesis that enlargements in size are inversely correlated with the mitochondrial metabolic competence.     

Ancient specimens and DNA contamination: a case study from the 12S rRNA gene sequence of the "Linh Duong" bovid (Pseudonovibos spiralis)

In 1993, several unusual horn sheaths, collected in the South of Vietnam, were regarded as evidence for a new mammal species, Pseudonovibos spiralis. However, the taxonomic status of P. spiralis remains a highly controversial subject: firstly, it has been related to three different groups of the family Bovidae: Antilopini (gazelles), Bovini (oxen, bisons, and buffaloes), and Caprini sensu lato (goats, sheep and allies). Secondly, certain horns have been shown to be faked. Most recently, it has been suggested, on the basis of 12S rRNA gene sequences, that P. spiralis was a new species of buffalo. I demonstrate here that this conclusion was inaccurate, and base this on the grounds that: (1) the putative sequence of P. spiralis is shown to be a chimera obtained from three different species: Bos taurus, Bubalus bubalis and Saiga tatarica, and (2) several factors indicate that the specimen used was not authentic. This new study confirms earlier suggestions that the horns used to formally describe P. spiralis were bogus and were in fact derived from the horns of domesticated cattle (Bos taurus). The data suggest that P. spiralis never existed.