Wing Sacs Research Articles

Bats, Bacteria, and Bat Smell V.2.0: Repeatable Sex-Specific Differences in Scent Organ Microbiota

Reproducibility is a fundamental principle in science, ensuring reliable and valid findings. However, replication studies are scarce, particularly in ecology, due to the emphasis on novelty for publication. We explored the possibility of replicating original findings in the field of microbial and chemical ecology by conducting a conceptual replication of a previous study analysing the sex-specific differences in the microbial communities inhabiting the wing sacs, a scent organ with crucial functions in olfactory communication, of greater sac-winged bat (Saccopteryx bilineata). In the original study, the skin swabs from the antebrachial wing sacs of the males and wing sac rudiments of the females were analysed using culture-dependent methods to test sex-specific differences. The authors demonstrated that males have lower microbial richness and different microbial composition than females. We attempted to reproduce these findings using 16S rRNA sequencing, which offers improved accuracy in pinpointing microbial members than culture-dependent methods because of advanced statistical methods. Our study validated the original study’s findings: Males had a lower microbial richness, and the community composition differed between the sexes. Furthermore, in the current study, males had an increased abundance of bacteria that might potentially be involved in odour production and degradation of malodorous substances and antimicrobial production. Our conceptual replication study corroborated that microbes can play a role in shaping their host’s olfactory phenotype and consequently influence sexual selection. Furthermore, the current study emphasises the importance of replication efforts and hopefully encourages a culture that values replication studies in scientific practice.

Evidence of female preference for odor of distant over local males in a bat with female dispersal

AbstractGeographic variation of sexually selected male traits is common in animals. Female choice also varies geographically and several studies found female preference for local males, which is assumed to lead to local adaptation and, therefore, increases fitness. As females are the nondispersing sex in most mammalian taxa, this preference for local males might be explained by the learning of male characteristics. Studies on the preference of females in female-dispersing species are lacking so far. To find out whether such females would also show preferences for local males, we conducted a study on greater sac-winged bats (Saccopteryx bilineata), a species where females disperse and males stay in their natal colony. Male greater sac-winged bats possess a wing pouch that is filled with odoriferous secretion and fanned toward females during courtship display. In a combination of chemical analysis and behavioral preference tests, we analyzed whether the composition of wing sac secretion varies between two geographically distinct populations (300 km), and whether females show a preference for local or distant male scent. Using gas chromatography, we found significant differences in the composition of the wing sac odors between the two geographically distinct populations. In addition, the behavioral preference experiments revealed that females of both populations preferred the scent of geographically distant males over local males. The wing sac odor might thus be used to guarantee optimal outbreeding when dispersing to a new colony. This is—to our knowledge—the first study on odor preference of females of a species with female-biased dispersal.

Expression of matrix metalloproteinase genes during basement membrane degradation in the metamorphosis of Bombyx mori

The present study was conducted to clarify the involvement of the basement membrane (BM) in insect metamorphosis through analysis of the expression profile of two types of metalloproteinase (MMP and ADAMTS) genes in several organs, their ecdysone involvement, and the histological change of BM. BM was observed around wing sac and in the wing cavity and around fat bodies at the W0 stage but disappeared after the W3 stage, and wing discs evaginated and fat body cells scattered after the W3 stage. The disappearance of the BM of midgut and silk glands was not observed after the W3 stage, but degenerated epithelium cells in the midgut and shrunken cells in the silk gland were observed after the W3 stage. BmMMP1 showed a peak at P0 in the wing discs, fat bodies, midgut, and silk gland. BmMMP2 showed a broad peak around pupation in the wing discs, fat bodies, midgut, and silk gland. BmADAMTS-1 showed enhanced expression at W2 in the wing discs, fat bodies, midgut, and hemocyte, while BmADAMTS-L showed enhanced expression at W3 in the fat bodies, midgut, silk gland, and hemocyte. After pupation, they showed a different expression in different organs. All of four genes were induced by 20-hydroxyecdysone in wing discs in vitro. The present results suggested the involvement of MMPs and ADAMTS in the BM digestion and the morphogenesis of organs during Bombyx metamorphosis.

First record of the rare bat Cyttarops alecto (Thomas, 1913) (Chiroptera: Emballonuridae) for the western Brazilian Amazonia, with comments on the type locality

Abstract Cyttarops alecto is a rare bat, poorly recorded along its distribu-tional range. In this paper, we report the fi rst record of C. alecto for the western Brazilian Amazon, provide information on dis-tribution and morphological variation of the species along with taxonomic comments based on comparisons of our specimen with the previous information reported in the literature, and we reinterpret the precise type locality for the species.Keywords: Brazil; distribution; emballonurids; Rond o nia; western Amazonia. The subtribe Diclidurina (Chiroptera: Emballonuridae) is confi ned to the Neotropical region and contains eight gen-era (Lim 2007 ). The monotypic genus and species Cyttarops consecutive nights near potential prey for vampire bats, such alecto (Thomas 1913 ), the smoky sheath-tailed short-eared bat, is medium-sized (forearm 45 – 47 mm) with a blackish gray, long, and silky dorsal pelage. Cyttarops has a greatly expanded clavicle, a deeply grooved tibia, and has no wing sac (Jones and Hood 1993 ). Cyttarops alecto is a rare bat, poorly recorded in its distri-butional range, and most captures of the species are of single individuals (Hood and Gardner 2007 , Velazco et al. 2011 ). Although documented by few specimens in collections, this species is broadly distributed in the Neotropical lowlands. The collecting localities of this species are distributed from Nicaragua to Panama, in Central America (Starrett and de and Langtimm 1993 , Jung et al. 2007 ), southwards in South America to eastern Colombia, northern Bolivia, and central Peru (Ochoa et al. 1994 , Aguirre et al. 2010 , Velazco et al. 2011 ) and eastwards to Guiana Shield localities in Guyana, French Guiana, and Suriname (Thomas 1913 , Masson and Cosson 1992 , Lim 2009 ) to the northern state of Par a , Brazil (the type locality), and southern Amazonian Brazil at the Tocantins River Basin (Nunes et al. 2006 ) (Figure 1 ). Herein, we report the fi rst record of

Scents of Adolescence: The Maturation of the Olfactory Phenotype in a Free-Ranging Mammal

Olfaction is an important sensory modality for mate recognition in many mammal species. Odorants provide information about the health status, genotype, dominance status and/or reproductive status. How and when odor profiles change during sexual maturation is, however often unclear, particularly in free-ranging mammals. Here, we investigated whether the wing sac odorant of male greater sac-winged bats (Saccopteryx bilineata, Emballonuridae) differs between young and adults, and thus offers information about sexual maturity to potential mating partners. Using gas chromatography – mass spectrometry, we found differences in the odorants of young and adult males prior and during, but not after the mating period. The wing sac odorant of adult males consists of several substances, such as Pyrocoll, 2,6,10-trimethyl-3-oxo-6,10-dodecadienolide, and a so far unidentified substance; all being absent in the odor profiles of juveniles prior to the mating season. During the mating season, these substances are present in most of the juvenile odorants, but still at lower quantities compared to the wing sac odorants of adults. These results suggest that the wing sac odorant of males encodes information about age and/or sexual maturity. Although female S. bilineata start to reproduce at the age of half a year, most males of the same age postpone the sexual maturation of their olfactory phenotype until after the first mating season.

Temporal and Spatial Distribution of Male Scent Marks in the Polygynous Greater Sac-Winged Bat

Scent marks are relatively long-lived signals that can be perceived by conspecifics when the producer is absent. Therefore, it is often not obvious to whom the signal is directed. In daytime roosts of the polygynous greater sac-winged bat, males scent mark territories with facial gland secretions. Territories are a valuable resource for males, as they offer exclusive courtship opportunities, which results in increased male reproductive success and, consequently, increased male-male competition over territories. The information encoded in male scent marks could, therefore, be either directed at females as part of an olfactory courtship display or at male competitors as part of territorial behaviour. We expected territorial males to scent mark in the morning, shortly before females return to the territory and close to female roosting sites, if scent marks are directed at females as part of the courtship display. And we expected harem males to scent mark at the territory boundaries, where male-male encounters are most likely to occur, if scent marks are directed at male competitors. We found that males marked more frequently in the afternoon, at a time when all females have already left the territory, and harem males marked at the territory boundaries and not inside their territory in the area where females roost. At boundaries males fan volatiles from specialised wing sacs towards competitors outside the territory. Scent marking of male Saccopteryx bilineata might therefore be congruent with the assessment-hypothesis, which states that scent marks offer intruders the possibility to make an olfactory assessment of the territory owner without direct physical interaction. Thus, scent marks of male S. bilineata are most likely influenced by male-male competition and not by female choice.

Odour-based species recognition in two sympatric species of sac-winged bats (Saccopteryx bilineata, S. leptura): combining chemical analyses, behavioural observations and odour preference tests

Combining chemical analysis and odour preference tests, we asked whether two closely related sympatric species of sac-winged bats use odour for species recognition. Males of the two sister species Saccopteryx bilineata and Saccopteryx leptura have pouches containing an odoriferous liquid in their antebrachial wing membrane, which is used in S. bilineata during courtship displays. Although both species occasionally share the same daytime roosts and are morphologically similar, there is no evidence for interbreeding. We compared the production and composition of the wing sac odorant in male S. leptura and S. bilineata and performed odour preference tests with female S. bilineata. Similar to male S. bilineata, male S. leptura cleansed and refilled their wing sacs with secretions, but they spent more time each day in doing so than male S. bilineata. Chemical analysis by gas chromatography and mass spectrometry revealed that male Saccopteryx carried species-specific scents in their wing sacs. Binary choice tests confirmed that female S. bilineata preferred the wing sac scents of male S. bilineata to those of the sister species, suggesting that the species specificity of male wing sac scents maintain the pre-mating isolation barrier between these closely related species.

Evolutionary Patterns of Morphology and Behavior as Inferred from a Molecular Phylogeny of New World Emballonurid Bats (Tribe Diclidurini)

A molecular phylogeny of New World emballonurid bats based on parsimony and Bayesian analyses of loci from the three different nuclear genetic transmission pathways in mammals (autosomal, X, and Y chromosomes) is well supported and independently corroborated by each individual gene tree. This is in contrast to a single most parsimonious but poorly supported tree based on morphological data, which has only one intergeneric or higher relationship shared with the molecular phylogeny. Combining the morphological and molecular data partitions results in a tree similar to the molecular tree suggesting a high degree of homoplasy and low phylogenetic signal in the morphological data set. Behavioral data are largely incomplete and likewise produce a poorly resolved tree. Nonetheless, patterns of evolution in morphology and behavior can be investigated by using the molecular tree as a phylogenetic framework. Character optimization of the appearance of dorsal fur and preferred roosting sites maps consistently and are correlated on the phylogeny. This suggests an association of camouflage for bats with unusual appearance (two dorsal stripes in Rhynchonycteris and Saccopteryx, or pale fur in Cyttarops and Diclidurus) and roosting in exposed sites (tree trunks or under palm leaves). In contrast, the ancestral states for Old and New World emballonurids are typically uniform brown or black, and they usually roost in sheltered roosts such as caves and tree hollows. Emballonuridae is the only family of bats that has a sac-like structure in the wing propatagium, which is found in four New World genera. Mapping the wing sac character states onto the phylogeny indicates that wing sacs evolved independently within each genus and that there may be a phylogenetic predisposition for this structure. Ear orientation maps relatively consistently on the molecular phylogeny and is correlated to echolocation call parameters and foraging behavior, suggesting a phylogenetic basis for these character systems.

BATS, BACTERIA, AND BAT SMELL: SEX-SPECIFIC DIVERSITY OF MICROBES IN A SEXUALLY SELECTED SCENT ORGAN

Microbes interfere with the olfactory communication of animals by degrading chemical signals or by adding volatile metabolites. We report on the composition and diversity of the microflora in a sexually selected scent organ, the wing sacs of Saccopteryx bilineata, which are used by males for courting females. Wing sacs lack any glandular tissues. Instead, males clean and refill their wing sacs each day with genital and gular secretions. Females have only a nonfunctional rudiment of this organ. We isolated a total of 40 microbial species with only a moderate overlap in species composition between the sexes. The estimated microbial diversity was significantly lower in males than in females, with a minimum of 52.5 microbial species ± 5.0 SD in wing sac rudiments of females and 40.3 ± 4.2 SD in wing sacs of males (jackknife estimates). Males carried on average only 2 out of 40 possible microbial species in their wing sacs. Thus, individual scent profiles of males could originate from individual microflora. The daily routine of wing sac cleaning and refilling has possibly evolved to control microbial scent degradation, to support an individual microflora involved via volatile metabolites in mate choice, or both. Microbes may play a more prominent role in the evolution of morphological structures and behavioral adaptations than previously envisaged.

Individual variation in perfume blending in male greater sac-winged bats

Male greater sac-winged bats, Saccopteryx bilineata, use hovering flights to court females in their harem territory. While hovering, males fan a fragrant perfume from sac-like organs in the wing membrane towards the females. Each afternoon, males renew the perfumes of their wing sacs during a stereotypical and time-consuming behavioural sequence, which includes blending secretions from genital and gular glands. I investigated whether male perfume-blending behaviour varies between the mating and the nonmating season, and whether successful males, those with large harems, have different patterns of perfume blending than less successful males. I measured variation in perfume-blending behaviour in 21 adult males. The pattern of perfume blending was not significantly different between the mating and the nonmating season. However, the time at which perfume blending began and terminated differed between seasons. During the mating season, males spent about the same time cleaning their wing sacs during phase I of perfume blending, irrespective of the number of females in their harem. During phase II, males with large harems spent significantly less time refilling and blending perfume than males with small harems. In addition, males with large harems took up fewer droplets of secretion from the genital region. Overall, male–male interactions were rare during perfume blending, and the decrease in duration in phase II was not attributable to more disturbance in large harems. There was also no evidence that males prevented each other from transferring secretions into their wing sacs. Copyright 2002 The Association for the Study of Animal Behaviour. Published by Elsevier Science Ltd. All rights reserved.

A histological examination of the holding sacs and glandular scent organs of some bat species (Emballonuridae, Hipposideridae, Phyllostomidae, Vespertilionidae, and Molossidae)

Using histological techniques at the light-microscope level, we examined and compared structure and sexual dimorphism of the wing sacs and integumentary glandular scent organs of 11 species of microchiropteran bats. The antebrachial wing sacs of the Neotropical emballonurids Peropteryx macrotis, Saccopteryx bilineata, and Saccopteryx leptura differed in size and location but lacked sudoriferous and sebaceous glands, confirming that they were holding sacs rather than glandular scent organs. Glandular scent organs from 11 species consisted of sebaceous and (or) sudoriferous glands in emballonurids (P. macrotis, S. bilineata, S. leptura, Taphozous melanopogon, Taphozous nudiventris), hipposiderids (Hipposiderous fulvus, Hipposiderous ater), the phyllostomid Sturnira lilium, the vespertilionid Rhogeessa anaeus, and molossids (Molossus ater and Molossus sinaloe). Glandular scent organs were located on the face (H. fulvus, H. ater), gular region (S. bilineata, P. macrotis, T. melanopogon, M. ater, M. sinaloe), chest (T. nudiventris), shoulder (S. lilium), or ears (R. anaeus). Glandular scent organs showed greater similarities within than between families, and typically were rudimentary or lacking in females. Scanning electron microscope examination revealed that the hairs associated with glandular areas of male T. melanopogon were larger and had a different cuticular-scale pattern than body hairs. These were osmetrichia, hairs specialized for holding and dispersing glandular products. In S. lilium, hairs associated with the shoulder scent-gland area were larger than body hairs but similar in cuticular-scale pattern.

Storage and display of odour by male Saccopteryx bilineata (Chiroptera, Emballonuridae)

Males of the sac-winged bat, Saccopteryx bilineata, actively fill their propatagial sacs with secretions from the genital region, the gular gland, urine and saliva. From our observations and those of Starck we deduce that propatagial sacs in S. bilineata do not have a glandular function, but are instead organs for the storage and display of odours. In addition to the already known “salting” and hovering behaviour of male S. bilineata, we describe in detail how odour is fanned to roosting individuals during the complex, stereotypic hovering displays. S. bilineata males also coat the fur of their backs with saliva using the wing tip and might scent-mark territory boundaries. “Yawning” may represent a visual as well as an olfactory cue. Odour seems to play an important role in the social communication of S. bilineata and in other emballonurids, as revealed by the broad distribution of wing sacs in this family. S. bilineata males display odour during energetically costly hovering flights in front of females. We demonstrate that the number of hovering displays increases with harem size. The mating effort of S. bilineata males comprises a multimodal signalling behaviour. Although males defend harem territories in which females gather, females seem to be able to choose the father of their progeny freely among the males of a colony. This may have led to the evolution of the complex mating displays by male S. bilineata.

Sequence complexities of mRNA populations during adult wing development in the silkmoth Antheraea polyphemus

The total poly A + messenger RNAs of epidermal cells in the wing sac of the silkmoth Antheraea polyphemus as a function of development (which is under the influence of the moulting hormone, 20-hydroxyecdysone) has been analyzed by cDNA-mRNA hybridizations. Kinetic and numerical analysis of homologous hybridizations indicate the messenger populations to be made up of several kinetic classes, the total complexity being similar at all stages of development. Heterologous hybridizations indicate clear differences between two stages: (i) at the beginning of development characterized by growth and development and (ii) at the end of development characterized by differentiation and cuticular secretion. The principal difference between the two stages has been shown, by isolation and characterization of null cDNA (cDNA enriched for the second stage) to be due to the appearance of several hundred new mRNAs belonging to the moderately abundant class at the second stage. Analysis of translation products by SDS-PAGE and immunoprecipitation confirm and extend these results.