Gas-Chromatographic Analyses of the Subcaudal Gland Secretion of the European Badger (Meles meles) Part II: Time-Related Variation in the Individual-Specific Composition

Individuality in body odors has been described in a variety of species, but studies on time-related variation in individual scent are scarce. Here, we use GC-MS to investigate how chemical composition of subcaudal gland secretions of European badgers (Meles meles) varies over days, seasons, and from year to year, and how secretions change with the length of time for which they are exposed to the environment. Samples were divided into subsamples—one was frozen immediately and the remaining ones frozen after 2, 6, 12, 24, and 48 hr, respectively—and many individual-specific characteristics of the scent-profiles remained stable over time. However, two components were negatively correlated with time, thus providing the possibility to determine the age of scent marks. The low variation found in scent profiles of samples collected from the same individual three days apart showed that the individual-specific scent is a true characteristic of the respective badger, and that trapping and subsequent sampling have little effect on the composition of subcaudal gland secretions. Long-term variation (i.e., over one year) in individual subcaudal scent profiles is not continuous, but periods of relative stability are followed by periods of rapid change, that can be related to badger biology. Annual variation between samples collected from the same individuals in winter 1998 and winter 1999, and in spring 1998 and spring 1999 was lower than seasonal variation. Therefore, the results of this study indicate the potential of an individual-specific scent signature in the subcaudal gland secretions of badgers evidencing that individual recognition is of high importance in this species.     

Intra- and interindividual variation in flank gland secretions of free-ranging shrewsCrocidura russula

Individual differences in flank gland secretions were examined among males of the monogamous shrewCrocidura russula during the breeding and nonbreeding seasons. Gas chromatography was used to measure intra- and interindividual variation of flank gland secretions of free-ranging shrews from different populations. The number of compounds detected by gas Chromatographic analyses was correlated with body mass, flank gland size, and the presence of blood parasites in individual shrews. Very few compounds were detected from the flank gland area of juvenile males. After they reached sexual maturity, however, the number of compounds detected from the flank gland secretions increased significantly. At the beginning of the reproductive season 48 different compounds were detected from male flank gland secretions. In the middle of the breeding season 70 compounds were detected, while only 11 compounds were detected during the nonbreeding season. Few compounds were common to all males. There were more volatile compounds in the flank gland secretions of males in the beginning of the breeding season than later in the breeding season. Males from the same population had fewer differences in the elution profile of compounds than males from different populations indicating that individuals from a distinct population have similar elution profiles of compounds and that each population has its own type of elution profile. No correlations were found between the number of compounds detected by gas chromatography for each male and the male's body mass or flank gland size. Blood parasites (trypanosomes,Trypanosoma crocidurae) were found in only three of 30 males investigated.     

Anal gland secretion of European mole: Volatile constituents and significance in territorial maintenance

The chemical composition of the volatile components of the anal gland secretion of mature and immature moles of both sexes was examined by gas chromatography and mass spectrometry. The compounds in the secretion of adult males appear to vary little throughout the year and are similar to those from the adult female outside the breeding season. The secretions are dominated by C₅-C₁₀ carboxylic acids. Female glands regress as they enter proestrus, accompanied by profound changes in the chemical composition of the secretion with early disappearance of the carboxylic acids. In juvenile moles, the composition of the secretion changes as the animal matures, with carboxylic acids becoming dominant only as the animal reaches maturity. Anal gland secretion probably plays an important role in territorial maintenance.     

Variation in Preen Oil Composition Pertaining to Season,Sex, and Genotype in the Polymorphic White-Throated Sparrow

Evidence for the the ability of birds to detect olfactory signals is now well documented, yet it remains unclear whether birds secrete chemicals that can be used as social cues. A potential source of chemical cues in birds is the secretion from the uropygial gland, or preen gland, which is thought to waterproof, maintain, and protect feathers from ectoparasites. However, it is possible that preen oil also may be used for individual recognition, mate choice, and signalling social/sexual status. If preen oil secretions can be used as socio-olfactory signals, we should be able to identify the volatile components that could make the secretions more detectable, determine the seasonality of these secretions, and determine whether olfactory signals differ among relevant social groups. We examined the seasonal differences in volatile compounds of the preen oil of captive white-throated sparrows, Zonotrichia albicollis. This species is polymorphic and has genetically determined morphs that occur in both sexes. Mating is almost exclusively disassortative with respect to morph, suggesting strong mate choice. By sampling the preen oil from captive birds in breeding and non-breeding conditions, we identified candidate chemical signals that varied according to season, sex, morph, and species. Linear alcohols with a 10–18 carbon chains, as well as methyl ketones and carboxylic acids, were the most abundant volatile compounds. Both the variety and abundances of some of these compounds were different between the sexes and morphs, with one morph secreting more volatile compounds in the non-breeding season than the other. In addition, 12 compounds were seasonally elevated in amount, and were secreted in high amounts in males. Finally, we found that preen oil signatures tended to be species-specific, with white-throated sparrows differing from the closely related Junco in the abundances and/or prevalence of at least three compounds. Our data suggest roles for preen oil secretions and avian olfaction in both non-social as well as social interactions.     

The Effect of Caste and Reproductive State on the Chemistry of the Cephalic Labial Glands Secretion of Bombus Terrestris

The cephalic labial glands are well developed in many bee species. In bumble bee males, they cover most of the head volume, and their secretion is used in marking reproductive territories and attracting virgin queens. In females, however, they are poorly studied. Here, we present chemical analyses of their secretion in queens and workers of Bombus terrestris under various social conditions. The secretion revealed a context-dependent composition with sterile females possessing large amounts of fatty acid dodecyl esters, ranging from dodecyl hexanoate to dodecyl oleate, compared to small amounts in fertile females. Significant reduction in the dodecyl esters also was found in queens at the competition phase, where worker reproduction, aggression, and gyne differentiation occur. The exclusive production of esters by sterile individuals also is typical of Dufour’s gland secretion in this species, albeit in this case these are octyl rather than dodecyl esters, and the differences between sterile and fertile individuals are qualitative rather than quantitative. We propose that the labial gland esters provide yet another signal of reproductive inactivity. In virgin queens, it may signal that egg laying is deferred to the next season, while in workers it reinforces the message “I am sterile and out of the reproductive competition”. Whether the reduction in dodecyl esters in fertile queens as a function of colony social development is involved in regulating gyne production and the onset of the competition phase is yet to be deciphered.     

Chemical Defense in Harvestmen (Arachnida,Opiliones): Do Benzoquinone Secretions Deter Invertebrate and Vertebrate Predators?

Two alkylated 1,4-benzoquinones were identified from the defensive secretion produced by the neotropical harvestman Goniosoma longipes (Gonyleptidae). They were characterized as 2,3-dimethyl-1,4-benzoquinone and 2-ethyl-3-methyl-1,4-benzoquinone. We tested the effectiveness of these benzoquinone secretions against several predator types, including invertebrates and vertebrates. Different predators were exposed to the harvestmen's gland secretion or to distilled water in laboratory bioassays. Our results indicate that secretions containing the 1,4-benzoquinones released by G. longipes can be an effective defense against predation, and that the effectiveness of the secretion is dependent on the predator type. The scent gland secretion repelled seven ant species, two species of large wandering spiders, and one frog species, but was not an effective defense against an opossum. Our study also demonstrates that the scent gland secretion of G. longipes can work as a chemical shield preventing the approach of three large predatory ants for at least 10 min. The chemical shield may protect the harvestman against successive attacks of the same ant worker and also allow the harvestman to flee before massive ant recruitment. Our data support the suggestion that chemical defenses may increase survival with some but not all potential predators. This variation in defense effectiveness may result from many interacting factors, including the attack strategy, size, learning ability, and physiology of the predators, as well as the chemical nature of the defensive compounds, type of emission, and amount of effluent released by the prey.     

Physiological sources of variation in chemical defense ofOreina gloriosa (Coleoptera: Chrysomelidae)

The defensive secretion of the alpine chrysomelidOreina gloriosa is a complex mixture of mainly cardenolides and tyrosine betaine. Individually sampled secretions of adult laboratory-reared and field-collected beetles were analyzed by reverse-phase HPLC; 16 secretion components were quantified. Quantities and concentrations of different components were significantly affected by the age, sex, and reproductive status of individual beetles. Aging was correlated with marked increases (up to 4.4-fold) and decreases (up to 2.7-fold) of quantities and concentrations of several components. Differences between the sexes were smaller, but quantities of all components and concentrations of several components were larger in laboratory-reared females than in males. There was less of one component of the secretion in mated than unmated females, but the concentrations of four secretion components were higher (up to 1.6-fold) in mated females.     

Chemistry of male mandibular gland secretions ofPhilanthus triangulum

Males of the European and African beewolf,Philanthus triangulum, possess a sex specific mandibular gland secretion that is used for marking plant stems in mating territories. The secretion is composed of 90% (Z)-11-eicosen-1-ol plus small amounts of 10-nonadecen-2-one, nonadecenal, octadecanoic and octadecenoic acids, and eicosenal. The chemistry of this secretion differs markedly from the secretions of North AmericanPhilanthus, which consist of a larger number of components that possess different chemical functionalities and are more volatile. We postulate that the chemical differences betweenP. triangulum and its New World relatives reflect phylogenetic differences plus a possible reduced necessity for species isolating mechanisms inP. triangulum.     

Chemical Characterization of Young Virgin Queens and Mated Egg-Laying Queens in the Ant <Emphasis Type="Italic">Cataglyphis cursor</Emphasis>: Random Forest Classification Analysis for Multivariate Datasets

Social insects are well known for their extremely rich chemical communication, yet their sex pheromones remain poorly studied. In the thermophilic and thelytokous ant, Cataglyphis cursor, we analyzed the cuticular hydrocarbon profiles and Dufour’s gland contents of queens of different age and reproductive status (sexually immature gynes, sexually mature gynes, mated and egg-laying queens) and of workers. Random forest classification analyses showed that the four groups of individuals were well separated for both chemical sources, except mature gynes that clustered with queens for cuticular hydrocarbons and with immature gynes for Dufour’s gland secretions. Analyses carried out with two groups of females only allowed identification of candidate chemicals for queen signal and for sexual attractant. In particular, gynes produced more undecane in the Dufour’s gland. This chemical is both the sex pheromone and the alarm pheromone of the ant Formica lugubris. It may therefore act as sex pheromone in C. cursor, and/or be involved in the restoration of monogyny that occurs rapidly following colony fission. Indeed, new colonies often start with several gynes and all but one are rapidly culled by workers, and this process likely involves chemical signals between gynes and workers. These findings open novel opportunities for experimental studies of inclusive mate choice and queen choice in C. cursor.     

The Role of Diet in Shaping the Chemical Signal Design of Lacertid Lizards

Lizards communicate with others via chemical signals, the composition of which may vary among species. Although the selective pressures and constraints affecting chemical signal diversity at the species level remain poorly understood, the possible role of diet has been largely neglected. The chemical signals of many lizards originate from the femoral glands that exude a mixture of semiochemicals, and may be used in a variety of contexts. We analyzed the lipophilic fraction of the glandular secretions of 45 species of lacertid lizard species by gas chromatography/mass spectrometry. The proportions of nine major chemical classes (alcohols, aldehydes, fatty acids, furanones, ketones, steroids, terpenoids, tocopherols and waxy esters), the relative contributions of these different classes (‘chemical diversity’), and the total number of different lipophilic compounds (‘chemical richness’) varied greatly among species. We examined whether interspecific differences in these chemical variables could be coupled to interspecific variation in diet using data from the literature. In addition, we compared chemical signal composition among species that almost never, occasionally, or often eat plant material. We found little support for the hypothesis that the chemical profile of a given species’ secretion depends on the type of food consumed. Diet breadth did not correlate with chemical diversity or richness. The amount of plants or ants consumed did not affect the relative contribution of any of the nine major chemical classes to the secretion. Chemical diversity did not differ among lizards with different levels of plant consumption; however, chemical richness was low in species with an exclusive arthropod diet, suggesting that incorporating plants in the diet enables lizards to increase the number of compounds allocated to secretions, likely because a (partly) herbivorous diet allows them to include compounds of plant origin that are unavailable in animal prey. Still, overall, diet appears a relatively poor predictor of interspecific differences in the broad chemical profiles of secretions of lacertid lizards.     

Volatile and Semivolatile Compounds in Gray Catbird Uropygial Secretions Vary with Age and Between Breeding and Wintering Grounds

The uropygial secretions of some bird species contain volatile and semivolatile compounds that are hypothesized to serve as chemical signals. The abundance of secretion components varies with age and season, although these effects have not been investigated in many species. We used solid-phase microextraction headspace sampling and solvent extraction coupled with gas chromatography–mass spectrometry to detect and identify volatile and semivolatile chemical compounds in uropygial secretions of gray catbirds (Dumetella carolinensis). We identified linear and branched saturated carboxylic acids from acetic (C2) through hexacosanoic (C26); linear alcohols from decanol (C10) through docosanol (C22); one aromatic aldehyde; one monounsaturated carboxylic acid; two methyl ketones; and a C28 ester. We tested for the effect of age on signal strength and found that juvenile birds produced greater amounts of volatile C4 through C7 acids and semivolatile C20 through C26 acids, although the variation among individuals was large. Adult birds displayed small concentrations and minimal individual variation among volatile compounds, but produced significantly higher levels of long-chain linear alcohols than juvenile birds. We tested for the effects of season/location by sampling adult catbirds at their Ohio breeding grounds and at their Florida wintering grounds and found that the heaviest carboxylic acids are significantly more abundant in secretions from birds sampled during winter at the Florida site, whereas methyl ketones are more abundant in birds sampled during summer on the Ohio breeding grounds. We observed no effect of sex on semivolatile compounds, but we found a significant effect of sex on levels of carboxylic acids (C4 through C7) for juvenile birds only.     

Use of Chemical Communication by the Subterranean Rodent Ctenomys talarum (Tuco-Tuco) During the Breeding Season

Solitary subterranean rodents with a low frequency of direct contact between conspecifics are expected to use chemical communication to coordinate social and reproductive behavior. We examined whether reproductive tuco-tucos (Ctenomys talarum) were able to discriminate the reproductive condition, sex, and source population of conspecifics by means of chemical cues contained in urine, feces, soiled shavings, or anogenital secretions. During preference tests in which animals had direct contact with these chemical cues, tuco-tucos were able to determine the reproductive condition of opposite sex conspecifics independent of the source of odor. When only olfactory cues were available, both sexes discriminated reproductive condition of opposite sex individuals using urine. Females were also able to discriminate the reproductive condition of males using soiled shavings. Females spent more time investigating male odors than female odors; except in the case of feces, breeding males spent similar amounts of time investigating male and female odors. No preferences were detected for opposite sex urine from members of an animal's own versus another population. The role of chemical cues in territory defense and breeding performance by this highly territorial subterranean rodent is discussed.     

Male Scent Gland Signals Mating Status in Greater Spear-Nosed Bats, <Emphasis Type="Italic">Phyllostomus hastatus</Emphasis>

Chemical signals are ubiquitous, but often overlooked as potentially important for conveying information relevant for sexual selection. The male greater spear-nosed bat, Phyllostomus hastatus, possesses a sexually dimorphic gland on the chest that produces an odoriferous secretion. Here, we investigate the potential for this glandular secretion to act as a sexually selected olfactory signal by examining gland activity in and out of the mating season and determining if variation in its chemical composition reflects variation in male mating status or attributes of the individual. Based on gas chromatography-mass spectrometry (GC-MS) measurements of samples collected from wild bats roosting in caves in Trinidad, West Indies, we find that males that defend and roost with groups of females (harem holders) have significantly different chemical profiles from males found roosting in all male groups (bachelors). Additionally, profiles differed significantly among individuals. Taken together, these results suggest that this chemical signal has the potential to communicate both mating status and individual identity and thus could be used to mediate interactions among individuals within this harem-based social mating system.     

Metapleural Gland Secretion of the Leaf-cutter Ant Acromyrmex octospinosus: New Compounds and Their Functional Significance

Ants of the myrmicine tribe Attini live in symbiosis with a fungus that provides them with food. In return the ants maintain optimal growth conditions for the fungus, weed out competing microorganisms, and inhibit the growth conditions of these competitors by chemical means. We present a comprehensive analysis of metapleural gland secretions of Acromyrmex octospinosus, using a recently developed method for the analysis of polar compounds by gas chromatography and mass spectrometry. We show that the chemical identity and quantitative recovery of different compounds in the metapleural gland secretion depends upon the method used and the type of colony from which the samples are taken. In addition to the two compounds previously recorded in the metapleural gland secretion of Acromyrmex ants (indolacetic acid and myrmicacin), 20 new compounds were detected in the secretion of a random sample of workers from two laboratory colonies and two field colonies. These compounds span the whole range of carboxylic acids from acetic acid to the long-chain fatty acids but comprise also some alcohols, lactones, and keto acids. The possible function of this highly complex secretion mixture is discussed.     

Response of Male Domestic Rabbits (Oryctolagus cuniculus) to Inguinal Gland Secretion from Intact and Ovariectomized Females

European rabbits (Oryctolagus cuniculus) of both sexes possess a variety of skin glands important in chemical communication. These include sizable inguinal glands, the strong-smelling secretion from which accumulates in conspicuous pouches of skin on either side of the genital opening. To test the hypothesis that this secretion advertises sexual receptivity in females, inguinal gland secretions from nine intact and six ovariectomized females were presented on dummy rabbits or in a simultaneous choice paradigm, and the responses of nine stud males to these secretions were recorded. Although the males clearly perceived the stimuli, no differences in responses to secretions from the intact and ovariectomized does were observed, and no differences were found in the size, weight, or secretory activity of the does' glands. It is concluded that inguinal gland secretion plays little role in communicating sexual receptivity in female rabbits and that ovarian steroids play little part in regulating inguinal gland activity.     

Naphthoquinones and Anthraquinones from Scent Glands of a Dyspnoid Harvestman, <Emphasis Type="Italic">Paranemastoma quadripunctatum</Emphasis>

Extracts of Paranemastoma quadripunctatum (Opiliones, Dyspnoi, Nemastomatidae) contained seven components, all of which likely originated from the secretion of well-developed prosomal scent glands. The two main components (together accounting for more than 90% of the secretion) were identified as 1,4-naphthoquinone and 6-methyl-1,4-naphthoquinone. The minor components were 1,4-naphthalenediol, two methoxy-naphthoquinones (2-methoxy-1,4-naphthoquinone, and 2-methoxy-6-methyl-1,4-naphthoquinone) and two anthraquinones (2-methyl-9,10-anthraquinone and a dimethyl-9,10-anthraquinone). While some chemical data on scent gland secretions of the other suborders of Opiliones (Cyphophthalmi, palpatorean Eupnoi, and Laniatores) already exist, this is the first report on the scent gland chemistry in the Dyspnoi. Naphthoquinones are known scent gland exudates of Cyphophthalmi and certain Eupnoi, methoxy-naphthoquinones and anthraquinones are new for opilionid scent gland secretions.     

Regeneration of Complex Oil-Gland Secretions and Its Importance for Chemical Defense in an Oribatid Mite

Most oribatid mites possess a pair of opisthonotal exocrine glands that produce mostly complex, species-specific secretions. Such blends may contain more than 10 different compounds, but hardly anything is known about their primary biosynthesis or regeneration. I analyzed recovery of the 6 main components from the 11-compound secretion of the oribatid mite Archegozetes longisetosus Aoki, including the main chemical classes hydrocarbons, aromatics, and terpenes, during a 20-day time course after complete gland depletion. About 10?% of the original total secretion amount was restored after 24?hr, and after 2–6?days, the amount had reached the range of total amount observed in the control group. Most compounds were recovered at similar rates within the first 48?hr. An important exception was pentadecane, which was predominantly produced in the first few hours, suggesting that this compound is the main solvent of the secretion. Although relative amounts of the main compounds differed significantly over time, the complex profile of the whole secretion was stable and not confidently distinguishable among the sampling dates. The general recovery rate was high during the first 48?hr, about 25 times higher than in the remaining 18?days. The biological importance of this high initial investment was supported by predation experiments: the predacious rove beetle Stenus juno was first repelled after 48?hr when at least 25?% of secretions was restored.     

Lipid compounds in secretions of fishing bat,Noctilio leporinus (Chiroptera: Noctilionidae)

The distinctive odor ofNoctilio leporinus arises from oily secretions found beneath the wings in the subaxillary region. Analysis of secretions by gas chromatography-mass spectrometry identified 372 lipid compounds. Differences in number and chemical composition of glycolipids suggest that secretions of males from the same roost are more similar to each other than to other males or females. Differences in number and chemical composition of nonpolar lipids indicate that secretions of males are more similar to each other than to females. Since secretions differ between sexes, information on sexual identity and reproductive condition may be communicated. Individually unique lipid compositions further suggest that bats may be recognizable by their odor within the roost and while flying.     

Evolutionary Interactions Between Visual and Chemical Signals: Chemosignals Compensate for the Loss of a Visual Signal in Male <Emphasis Type="Italic">Sceloporus</Emphasis> Lizards

Animals rely on multimodal signals to obtain information from conspecifics through alternative sensory systems, and the evolutionary loss of a signal in one modality may lead to compensation through increased use of signals in an alternative modality. We investigated associations between chemical signaling and evolutionary loss of abdominal color patches in males of four species (two plain-bellied and two colorful-bellied) of Sceloporus lizards. We conducted field trials to compare behavioral responses of male lizards to swabs with femoral gland (FG) secretions from conspecific males and control swabs (clean paper). We also analyzed the volatile organic compound (VOC) composition of male FG secretions by stir bar extraction and gas chromatography-mass spectrometry (GC-MS) to test the hypothesis that loss of the visual signal is associated with elaboration of the chemical signal. Males of plain-bellied, but not colorful-bellied species exhibited different rates of visual displays when exposed to swabs of conspecific FG secretions relative to control swabs. The VOC composition of male Sceloporus FG secretions was similar across all four species, and no clear association between relative abundances of VOCs and evolutionary loss of abdominal color patches was observed. The emerging pattern is that behavioral responses to conspecific chemical signals are species- and context-specific in male Sceloporus, and compensatory changes in receivers, but not signalers may be involved in mediating increased responsiveness to chemical signals in males of plain-bellied species.     

Chemical communication in the primitive antAneuretus simoni: The role of the sternal and pygidial glands

Trail and alarm communication inAneuretus simoni are mediated by the secretions of the sternal and pygidial glands, respectively. The sternal gland is composed of a glandular epithelium and an associated reservoir located in the 7th sternum. This gland produces a relatively long-lived mass recruitment pheromone. The pygidial gland opens between the 6th and 7th tergites and produces a secretion that releases aggressive alarm. The evolution of glandular physiology in the Aneuretinae and Dolichoderinae are discussed in view of these findings.