Rousettus Leschenaulti Research Articles

Genetic diversity within and among populations of the fulvous fruit bat Rousettus leschenaulti using RAPD analysis

Genetic diversity within and among populations of the fulvous fruit bat Rousettus leschenaulti using RAPD analysis

Inward collapse of the nasal cavity: Perinatal consolidation of the midface and cranial base in primates.

Living primates show a complex trend in reduction of nasal cavity spaces and structures due to moderate to severe constraint on interorbital breadth. Here we describe the ontogeny of the posterior end of the primate cartilaginous nasal capsule, the thimble shaped posterior nasal cupula (PNC), which surrounds the hind end of the olfactory region. We used a histologically sectioned sample of strepsirrhine primates and two non-primates (Tupaia belangeri, Rousettus leschenaulti), and histochemical and immunohistochemical methods to study the PNC in a perinatal sample. At birth, most strepsirrhines possess only fragments of PNC, and these lack a perichondrium. Fetal specimens of several species reveal a more complete PNC, but the cartilage exhibits uneven or weak reactivity to type II collagen antibodies. Moreover, there is relatively less matrix than in the septal cartilage, resulting in clustering of chondrocytes, some of which are in direct contact with adjacent connective tissues. In one primate (Varecia spp.) and both non-primates, the PNC has a perichondrium at birth. In older, infant Varecia and Rousettus, the perichondrium of the PNC is absent, and PNC fragmentation at its posterior pole has occurred in the former. Loss of the perichondrium for the PNC appears to precede resorption of the posterior end of the nasal capsule. These results suggest that the consolidation of the basicranial and facial skeletons happens ontogenetically earlier in primates than other mammals. We hypothesize that early loss of cartilage at the sphenoethmoidal articulation limits chondral mechanisms for nasal complexity, such as interstitial expansion or endochondral ossification.

Responses of Short-Nosed Fruit Bat, <i>Cynopterus sphinx</i> (Vahl 1797) Towards Distress Calls of their Conspecifics from Related and Unrelated Sites:Implications for Building a Social Relationship

Distress calls emitted by bats signal their conspecifics either to warn them or inform them about the situations. Conspecifics may also get attracted towards distress calls as a behaviour of cooperative mobbing or just selfishly assessing the potential source of danger. The exact functions of distress calls in bats therefore vary to a great degree and are very hard to pinpoint. We conducted playback experiments to test the response of short-nosed fruit bat, Cynopterus sphinx towards the distress calls of their conspecifics from related and unrelated sites. Bats were attracted to their conspecifics from both related and unrelated sites and in one occasion towards fruit bat ( Rousettus leschenaulti ) of another genus within the same family. The response towards the opposite sex was significant in most of the playback trials and the reasons remain unclear. This symmetric response towards conspecifics from related and unrelated sites suggests the possibility of fruit bats building social relationships among unrelated individuals and probably between species.

Ultrastructure of the Secretory Epithelial Cells of the Bulbo-Urethral Glands (Cowper's gland) in the Indian Fruit Bat, Rousettus leschenaulti (Desmarest) During the Reproductive Cycle

Ultrastructure of the Secretory Epithelial Cells of the Bulbo-Urethral Glands (Cowper's gland) in the Indian Fruit Bat, Rousettus leschenaulti (Desmarest) During the Reproductive Cycle

A Bat-Derived Putative Cross-Family Recombinant Coronavirus with a Reovirus Gene.

The emergence of severe acute respiratory syndrome coronavirus (SARS-CoV) in 2002 and Middle East respiratory syndrome coronavirus (MERS-CoV) in 2012 has generated enormous interest in the biodiversity, genomics and cross-species transmission potential of coronaviruses, especially those from bats, the second most speciose order of mammals. Herein, we identified a novel coronavirus, provisionally designated Rousettus bat coronavirus GCCDC1 (Ro-BatCoV GCCDC1), in the rectal swab samples of Rousettus leschenaulti bats by using pan-coronavirus RT-PCR and next-generation sequencing. Although the virus is similar to Rousettus bat coronavirus HKU9 (Ro-BatCoV HKU9) in genome characteristics, it is sufficiently distinct to be classified as a new species according to the criteria defined by the International Committee of Taxonomy of Viruses (ICTV). More striking was that Ro-BatCoV GCCDC1 contained a unique gene integrated into the 3’-end of the genome that has no homologs in any known coronavirus, but which sequence and phylogeny analyses indicated most likely originated from the p10 gene of a bat orthoreovirus. Subgenomic mRNA and cellular-level observations demonstrated that the p10 gene is functional and induces the formation of cell syncytia. Therefore, here we report a putative heterologous inter-family recombination event between a single-stranded, positive-sense RNA virus and a double-stranded segmented RNA virus, providing insights into the fundamental mechanisms of viral evolution.

Cellular composition of the Corpus luteum of Indian fruit bat, Rousettus leschenaulti (Desmarest) during early embryonic development

The fine structure of luteal cell of the corpus luteum of Indian fruit bat, Rousettus leschenaulti was studied at three stages unilaminar blastocyst stage, Implanted bilaminar blastocyst stage and limb bud stage of early pregnancy. At unilaminar blastocyst stage luteal cells had small nuclei euchromatin. Mitochondria were small, round shaped with tubular cristae. Numerous less osmiophilic lipid droplets were observed in cytoplasmic field of the luteal cells. After implantation at implanted bilaminar blastocyst stage nuclear heterochromatin were reduced and nucleoli were larger and complex. Mitochondria were enlarged and often bizarre shaped with tubular cristae. Golgi complex and agranular endoplasmic reticulum were more conspicuous. Lipid droplets were less osmiophilic. At the stage of limb bud formation the luteal cells suggests different morphological picture, the nuclear size is reduced with clumps of heterochromatin. The agranular endoplasmic reticulum assumes the form of bundles of parallel tubules dispersed in several planes. Mitochondrial size was reduced then the previous stage and they posses vesicular cristae. These observations suggest that the steroidogenic activity of the luteal cells is highest during implantation and comparatively regresses during limb bud formation. It is suggested that the luteal cells is an important ovarian source of pregnancy hormones.

Ultrastructure of the secretory epithelial cells of the Cowper's gland in the Indian fruit bat, Rousettus leschenaulti (Desmarest) during the reproductive cycle

The present paper describes the ultrastructural characteristics of the Cowper’ glands of the Indian fruit bat, Rousettus leschenaulti during its sexually inactive-breeding cycle. The functional significance of the secretions of the Cowper's gland in reproduction is discussed. In Rousettus, Cowper's glands are small, pear-shaped, bilaterally symmetrical and are situated on either side of the base of the penis. Each lobule is made up of secretory acini lined by columnar or pyramidal cells. Ultrastructurally, the secretory epithelial cells are characterized by well-developed rough endoplasmic reticulum, extensively developed Golgi complex, and mitochondria. Three different types of secretory granules can be identified on the basis of electron density. These granules represent the different stages of granule maturation. The secretory products are released into the lumen both by apocrine and merocrine modes. The secretory material synthesized by the Cowper’ gland may be involved in various male reproductive processes of this species of bat.

Scanning electron microscopy study of guard hair of three Indian pteropodid bats

A comparative study on the hair microstructure of pteropodid bats Pteropus giganteus , Rousettus leschenaulti , and Cynopterus sphinx was performed using scanning electron microscope. Hair samples were taken from the dorsal, ventral, and neck regions. Among the three pteropodids examined, an imbricate and tightly appressed hair type was observed in P. giganteus , while a coronal type of cuticle with margins diverging from the shaft was observed in R. leschenaulti and C. sphinx . The coronal cuticles of R. leschenaulti and C. sphinx had relatively thinner scale widths than the imbricate cuticles of P. giganteus . There was a significant difference in hair length, scale lengths and scale widths, and scale indices among the three species of fruit bats. However, there was no significant difference in hair length and scale index of male and female R. leschenaulti as well as in hair length and scale indices of male and female C. sphinx . The presence of bell-shaped coronal cuticle and wide angular scale margin in R. leschenaulti and C. sphinx reveal their role as active pollinators. The hair patterns, hair length, scale length and width confirm the presence of species-specific characteristics of pteropodid bats and can be used for species identification.

Characterization of the mitochondrial genome of Rousettus leschenaulti

We present a complete sequence of mitochondrial genome of echolocating megabat Rousettus leschenaulti (16,704 nt, GenBank record KC702803) and provide its annotation. We also show that phylogenetic analysis based on mitochondrial genomes of eighteen bat species, including R. leschenaulti, supports division of Chiroptera into Yinpterochiroptera and Yangochiroptera.

Serological Evidence ofToxoplasma gondiiInfection in Five Species of Bats in China

Toxoplasma gondii is an obligate intracellular protozoan parasite that can infect almost all warm-blooded animals and humans with a worldwide distribution. Bats are reservoirs for an increasing number of emerging zoonotic viruses, such as henipaviruses and severe acute respiratory syndrome coronavirus (SARS-CoV). However, little is known of T. gondii infection in bats. The objective of the present study was to determine the seroprevalence of T. gondii infection in bats in China. A total of 217 serum samples from 5 species of bats were collected between April, 2010, and August, 2011, from 4 provinces in China. Antibodies to T. gondii were determined using the modified agglutination test (MAT, 1:25 or higher). Antibodies to T. gondii were found in 26.5% (18/68) Megaderma lyra, 13.6% (12/88) Rousettus leschenaulti, 13.6% (3/22) Cynopterus sphinx, 20% (4/20) Vespertilio superaus, and 15.8% (3/19) Pipistrellus javanicus. Antibody titers ranged from 1:25 to 1:400, with titers of 1:200 detected in 4 of the 5 bat species. The present study suggests the likely occurrence of T. gondii infection in bats in China, and these bats are new putative hosts for T. gondii, which may pose a threat to human health.

Recent Transmission of a Novel Alphacoronavirus, Bat Coronavirus HKU10, from Leschenault's Rousettes to Pomona Leaf-Nosed Bats: First Evidence of Interspecies Transmission of Coronavirus between Bats of Different Suborders

Although coronaviruses are known to infect various animals by adapting to new hosts, interspecies transmission events are still poorly understood. During a surveillance study from 2005 to 2010, a novel alphacoronavirus, BatCoV HKU10, was detected in two very different bat species, Ro-BatCoV HKU10 in Leschenault's rousettes (Rousettus leschenaulti) (fruit bats in the suborder Megachiroptera) in Guangdong and Hi-BatCoV HKU10 in Pomona leaf-nosed bats (Hipposideros pomona) (insectivorous bats in the suborder Microchiroptera) in Hong Kong. Although infected bats appeared to be healthy, Pomona leaf-nosed bats carrying Hi-BatCoV HKU10 had lower body weights than uninfected bats. To investigate possible interspecies transmission between the two bat species, the complete genomes of two Ro-BatCoV HKU10 and six Hi-BatCoV HKU10 strains were sequenced. Genome and phylogenetic analyses showed that Ro-BatCoV HKU10 and Hi-BatCoV HKU10 represented a novel alphacoronavirus species, sharing highly similar genomes except in the genes encoding spike proteins, which had only 60.5% amino acid identities. Evolution of the spike protein was also rapid in Hi-BatCoV HKU10 strains from 2005 to 2006 but stabilized thereafter. Molecular-clock analysis dated the most recent common ancestor of all BatCoV HKU10 strains to 1959 (highest posterior density regions at 95% [HPDs], 1886 to 2002) and that of Hi-BatCoV HKU10 to 1986 (HPDs, 1956 to 2004). The data suggested recent interspecies transmission from Leschenault's rousettes to Pomona leaf-nosed bats in southern China. Notably, the rapid adaptive genetic change in BatCoV HKU10 spike protein by ~40% amino acid divergence after recent interspecies transmission was even greater than the ~20% amino acid divergence between spike proteins of severe acute respiratory syndrome-related Rhinolophus bat coronavirus (SARSr-CoV) in bats and civets. This study provided the first evidence for interspecies transmission of coronavirus between bats of different suborders.

Serologic Evidence of Nipah Virus Infection in Bats, Vietnam

To the Editor: Bats are potential reservoir for highly pathogenic viruses, such as Nipah virus (NiV) and Hendra virus, which can cross species barriers (1). However, only limited surveillance has been conducted to assess risk for infection by these deadly emerging viruses. We conducted a study in Vietnam from 2007 to 2008 to assess the prevalence of these pathogens in bats. Different species of live bats were obtained from hunters or were captured in caves, pepper fields, and residential areas by using mist nets or harp traps (Technical Appendix Figure 1). A total of 451 serum samples were collected and subjected to IgG ELISA by using an Escherichia coli–derived recombinant nucleocapsid (N) protein of NiV (NiV-N-ELISA) (2). Two Leschenault’s rousette bats (Rousettus leschenaulti) were vaccinated with the same recombinant N protein to obtain positive serum specimens that contained antibodies against NiV-N. An optical density of 492 nm for negative control serum (1,000× dilution) was designated as 1:1,000 ELISA units. ELISA titers of sample serum specimens were obtained at a single dilution (1,000×) by using a standard curve of positive serum with high titers. A sample titer >3,000 was considered positive for IgG against NiV. Positive results were detected from only 2 fruit bat species, R. leschenaulti (31 bats [49.1%]) and Cynopterus sphinx (3 bats [2.8%]) (Table). Of the 34 samples positive by ELISA, only 22 (20 from the former and 2 from the latter bat species), which had enough volume left, were further analyzed by Western blot (WB) with E. coli–expressed recombinant N protein of NiV (Table; Technical Appendix Figure 2). Twenty of the 22 specimens were confirmed as positive by WB. ELISA-positive samples with high titers were also positive by WB for both bat species. However, only 1 sample from an R. leschenaulti bat was positive by WB that used a baculovirus-expressed recombinant N protein. Because of the different protein expression systems, the reactivity of bat antibody against NiV protein in WBs showed different patterns (Technical Appendix Figure 2). Neutralization tests (NTs) in which live NiVs (strain Ma-JMR-01–98) were used were performed at the Institute of Tropical Medicine, Nagasaki University. No specimens of C. sphinx bats were positive by NT; however, 2 specimens from R. leschenaulti bats, both positive with low titers, were confirmed to be positive by NT (50% cytopathic effect after NT, titers of 33.6 and 14.1). However, the latter specimen was negative by WB analysis. Table Results of serologic tests for Nipah virus on bats captured in Vietnam, 2007–2008* Seroepidemiologic studies in other countries have indicated that Pteropus spp. bats (fruit-eating bats) are the main reservoirs for NiV (3–6). Pteropid bats are usually found only in southern Vietnam. We could not obtain these bats for our study. However, a relatively high prevalence (49.1%) of henipavirus antibody was found in R. leschenaulti specimens from Hoa Binh Province. Rousettus spp. bats are the only megabats that use echolocation. These bats hang together on cave ceilings in a tightly packed manner and in groups composed of bats of both sexes and of different ages. They roost in large colonies and fly vast distances to find fruit (7). This behavior might be related to their high rate of seropositivity for viral infections. In southern China, bats of the same species showed a high prevalence of henipavirus antibody (8). R. leschenaulti bats are distributed from central to northern Vietnam and southern China. C. sphinx bats are common all over Vietnam, and their habitat overlaps with that of pteropid bats in southern Vietnam. Previous studies showed that IgG ELISA results for NiV-positive flying foxes correlated well with NT results (3,4). However, in our study, discrepancies existed between NT results and NiV-N-ELISA and WB results. A reason for these differences could be that Nipah-like viruses are circulating among bats in Vietnam, producing antibodies that are cross-reactive by ELISA and WB, but poorly cross-reactive by NT. The cross-reactive antibodies were probably not directed against neutralizing epitopes. To date, no reports have been made of an increased number of febrile encephalitis cases among the residents in Hoa Binh and Dak Lak Provinces where seropositive bats were captured. The circulating viruses may lack the pathogenic potential of Hendra and Nipah viruses. A survey by questionnaire was conducted among residents of Dak Nong and Dak Lak Provinces, where NiV-N-ELISA–positive C. sphinx bats were captured, to determine the frequency of contact between humans and bats. Risk factors for infection were observed in this study, such as bat hunting and cooking and drinking bat blood. In such situations, persons have direct contact with bat body fluids and feces and might be bitten during bat hunting. Thus, long-term systematic surveillance of bats is needed to determine the ecologic relationship between bats, humans, other animals, and the environment.

Seed dispersal in Hong Kong, China: past, present and possible futures

In the present article, published and unpublished information regarding seed dispersal in the degraded landscape of Hong Kong, China, is reviewed. Information was available for 1681 native plant species, of which 1165 were assigned to probable seed dispersal modes. Endozoochory accounted for one-third (34.4%) of all species evaluated, half (54.4%) of those from forests, and more than two-thirds (69.2%) of all trees and tall shrubs. Wind dispersal (25.9%) and dispersal by an unknown agent (30.7%) accounted for most of the rest, with the unknown species mostly small-seeded herbs. Although the frugivore fauna of the Hong Kong region has been truncated since the late Pleistocene, there are few clear examples of failed mutualisms. The most striking is the absence of scatter-hoarding rodents from Hong Kong, despite the presence of forest trees that appear to require them for effective dispersal. There are also some large Lauraceae fruits that appear to be targeted at larger-gaped birds than currently present. Most endozoochorous species are dispersed by 3 small passerine birds (Pycnonotus jocosus, P. sinensis, and Zosterops japonicus), with larger birds, fruit bats (Cynopterus sphinx, Rousettus leschenaulti), rhesus macaques (Macaca mulatta), civets (Paguma larvata, Viverricula indica), and muntjacs (Muntiacus muntjac) accounting for the remainder. The low plant diversity in frugivore droppings, seed traps, and secondary vegetation suggests plant succession may be dispersal limited, although this has not been investigated experimentally. Planting underdispersed species is the simplest solution but, in the longer term, the (re)introduction of a scatter-hoarding rodent should be considered as well as that of other locally and regionally extinct frugivores.

Contrasting genetic structure in two co-distributed species of old world fruit bat.

The fulvous fruit bat (Rousettus leschenaulti) and the greater short-nosed fruit bat (Cynopterus sphinx) are two abundant and widely co-distributed Old World fruit bats in Southeast and East Asia. The former species forms large colonies in caves while the latter roots in small groups in trees. To test whether these differences in social organization and roosting ecology are associated with contrasting patterns of gene flow, we used mtDNA and nuclear loci to characterize population genetic subdivision and phylogeographic histories in both species sampled from China, Vietnam and India. Our analyses from R. leschenaulti using both types of marker revealed little evidence of genetic structure across the study region. On the other hand, C. sphinx showed significant genetic mtDNA differentiation between the samples from India compared with China and Vietnam, as well as greater structuring of microsatellite genotypes within China. Demographic analyses indicated signatures of past rapid population expansion in both taxa, with more recent demographic growth in C. sphinx. Therefore, the relative genetic homogeneity in R. leschenaulti is unlikely to reflect past events. Instead we suggest that the absence of substructure in R. leschenaulti is a consequence of higher levels of gene flow among colonies, and that greater vagility in this species is an adaptation associated with cave roosting.

Bat Coronaviruses and Experimental Infection of Bats, the Philippines

Fifty-two bats captured during July 2008 in the Philippines were tested by reverse transcription-PCR to detect bat coronavirus (CoV) RNA. The overall prevalence of virus RNA was 55.8%. We found 2 groups of sequences that belonged to group 1 (genus Alphacoronavirus) and group 2 (genus Betacoronavirus) CoVs. Phylogenetic analysis of the RNA-dependent RNA polymerase gene showed that groups 1 and 2 CoVs were similar to Bat-CoV/China/A515/2005 (95% nt sequence identity) and Bat-CoV/HKU9-1/China/2007 (83% identity), respectively. To propagate group 2 CoVs obtained from a lesser dog-faced fruit bat (Cynopterus brachyotis), we administered intestine samples orally to Leschenault rousette bats (Rousettus leschenaulti) maintained in our laboratory. After virus replication in the bats was confirmed, an additional passage of the virus was made in Leschenault rousette bats, and bat pathogenesis was investigated. Fruit bats infected with virus did not show clinical signs of infection.

Foraging areas of Rousettus leschenaulti on the Hainan Island of China

Abstract We investigated the foraging area of three individuals (1 female and 2 males) of Rousettus leschenaulti (Chiroptera, Pteropodidae) in suburban Haikou City, Hainan Province, South China from November 2005 to January 2006 using radio telemetry. These animals left the daytime roosting sites about 90 min after sunset with no significant difference in departure time between the male and female bats. The average active times were 391.8 min for males and 533.7 min for the female, respectively. By reconstructing 93 radio-telemetry recording positions, we found that the long axis of foraging area of the bats ranged from 7.45 to 11.70 km. The foraging area of the female (3867 ha) was larger than that of the males (1138 ha), and there was overlap between the foraging areas of different individuals. These bats usually kept the same flight routes across a few successive days from the daytime roosting site to the foraging areas. Our findings suggested that female R. leschenaulti may explore a larger foraging area than males. No obvious territorial behaviors were observed in our studied area.

Seroprevalence of Rabies Virus Antibodies in Bats from Southern China

Members of the Order Chiroptera are the natural reservoirs of lyssaviruses and play an important role in the transmission of rabies to animals and humans. In this present study, the seroprevalence for rabies virus was determined for bats sampled from four southern provinces on the Chinese mainland. A total of 685 bats of 8 species representing 4 families were collected from 10 sites, and were tested by the indirect fluorescent antibody test using fluorescein isothiocyanate (FITC)-conjugated protein A/G mixture and viral neutralization test. Rabies antibody response was only detected from three bat species (Rousettus leschenaulti, Rhinolophus blythi, and Rhinolophus ferrumequinum). The overall rabies seroconversion rate was only 2.2% (15/685). Of the 15 positive sera, 13 (12 fruit bats and 1 insectivorous bat) were indirect fluorescent antibody test positive, and two insectivorous bats were virus neutralization positive when tested by the modified fluorescent antibody viral neutralization test, albeit extremely low. To our knowledge, this is the first published report describing rabies seroprevalences from Chinese bats. These results suggest that bats may play a role in the ecology of lyssaviruses in China, and further surveillance for the presence of lyssaviruses in bats should be undertaken throughout the country and extended to other species.

Host specificity in bat ectoparasites: A natural experiment

We undertook a field study to determine patterns of specialisation of ectoparasites in cave-dwelling bats in Sri Lanka. The hypothesis tested was that strict host specificity (monoxeny) could evolve through the development of differential species preferences through association with the different host groups. Three species of cave-dwelling bats were chosen to represent a wide range of host–parasite associations (monoxeny to polyxeny), and both sympatric and allopatric roosting assemblages. Of the eight caves selected, six caves were “allopatric” roosts where two of each housed only one of the three host species examined: Rousettus leschenaulti (Pteropodidae), Rhinolophus rouxi and Hipposideros speoris (Rhinolophidae). The remaining two caves were “sympatric” roosts and housed all three host species. Thirty bats of each species were examined for ectoparasites in each cave, which resulted in a collection of nycteribiid and streblid flies, an ischnopsyllid bat flea, argasid and ixodid ticks, and mites belonging to three families. The host specificity of bat parasites showed a trend to monoxeny in which 70% of the 30 species reported were monoxenous. Odds ratios derived from χ 2-tests revealed two levels of host preferences in less-specific parasites (i) the parasite was found on two host species under conditions of both host sympatry and host allopatry, with a preference for a single host in the case of host sympatry and (ii) the preference for a single host was very high, hence under conditions of host sympatry, it was confined to the preferred host only. However, under conditions of host allopatry, it utilized both hosts. There appears to be an increasing prevalence in host preferences of the parasites toward confinement to a single host species. The ecological isolation of the bat hosts and a long history of host–parasite co-existence could have contributed to an overall tendency of bat ectoparasites to become specialists, here reflected in the high percentage of monoxeny.

The role of olfaction and vision in the foraging behaviour of an echolocating megachiropteran fruit bat, Rousettus leschenaulti (Pteropodidae)

Many mammals use multimodal sensory information to find their food in complex environments. We studied the roles of olfactory and visual cues in the foraging behaviour of Rousettus leschenaulti, a tongue-clicking megachiropteran bat. We conducted experiments by offering a whole fruit, mashed fruit without shape and an artificial fruit to R. leschenaulti in dim light as well as in total darkness. R. leschenaulti responded to whole guava fruit and mashed guava fruit even in total darkness, whereas, artificial fruit similar to real fruit in size, shape and structure was ignored even when illuminated. These results reveal that odour may act as the primary cue for R. leschenaulti in finding fruit.

Karyotype evolution in Rhinolophus bats (Rhinolophidae, Chiroptera) illuminated by cross-species chromosome painting and G-banding comparison

Rhinolophus (Rhinolophidae) is the second most speciose genus in Chiroptera and has extensively diversified diploid chromosome numbers (from 2n = 28 to 62). In spite of many attempts to explore the karyotypic evolution of this genus, most studies have been based on conventional Giemsa staining rather than G-banding. Here we have made a whole set of chromosome-specific painting probes from flow-sorted chromosomes of Aselliscus stoliczkanus (Hipposideridae). These probes have been utilized to establish the first genome-wide homology maps among six Rhinolophus species with four different diploid chromosome numbers (2n = 36, 44, 58, and 62) and three species from other families: Rousettus leschenaulti (2n = 36, Pteropodidae), Hipposideros larvatus (2n = 32, Hipposideridae), and Myotis altarium (2n = 44, Vespertilionidae) by fluorescence in situ hybridization. To facilitate integration with published maps, human paints were also hybridized to A. stoliczkanus chromosomes. Our painting results substantiate the wide occurrence of whole-chromosome arm conservation in Rhinolophus bats and suggest that Robertsonian translocations of different combinations account for their karyotype differences. Parsimony analysis using chromosomal characters has provided some new insights into the Rhinolophus ancestral karyotype and phylogenetic relationships among these Rhinolophus species so far studied. In addition to Robertsonian translocations, our results suggest that whole-arm (reciprocal) translocations involving multiple non-homologous chromosomes as well could have been involved in the karyotypic evolution within Rhinolophus, in particular those bats with low and medium diploid numbers.