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Lyssavirus surveillance in bats was performed in Bangladesh during 2003 and 2004. No virus isolates were obtained. Three serum samples (all from Pteropus giganteus, n = 127) of 288 total serum samples, obtained from bats in 9 different taxa, neutralized lyssaviruses Aravan and Khujand. The infection occurs in bats in Bangladesh, but virus prevalence appears low.

Highlights

► Universal real-time PCR primer pair demonstrated to hybridize to and detect each of the known Lyssaviruses (including Rabies virus) with greater sensitivity than a standard pan-Lyssavirus hemi-nested RT-PCR typically used. ► Target sequences of bat derived virus species unavailable for analysis (Aravan-, Khujand-, Irkut-, West Caucasian bat- and Shimoni bat virus) were synthesized to produce oligonucleotides and the synthetic DNA was used as a target for primer hybridization.

Rabies virus (RABV) is enzootic throughout most of the world. It is now widely accepted that RABV had its origins in bats. Ten of the 11 Lyssavirus species recognised, including RABV, have been isolated from bats. There is, however, a lack of understanding regarding both the ecology and host reservoirs of Lyssaviruses. A real-time PCR assay for the detection of all Lyssaviruses using universal primers would be beneficial for Lyssavirus surveillance. It was shown that using SYBR® Green, a universal real-time PCR primer pair previously demonstrated to detect European bat Lyssaviruses 1 and 2, and RABV, was able to detect reverse transcribed RNA for each of the seven virus species available to us. Target sequences of bat derived virus species unavailable for analysis were synthesized to produce oligonucleotides. Lagos Bat-, Duvenhage- and Mokola virus full nucleoprotein gene clones enabled a limit of 5–50 plasmid copies to be detected. Five copies of each of the synthetic DNA oligonucleotides of Aravan-, Khujand-, Irkut-, West Caucasian bat- and Shimoni bat virus were detected. The single universal primer pair was therefore able to detect each of the most divergent known Lyssaviruses with great sensitivity.

Background and objectives

Bats are recognized as a major reservoir of lyssaviruses; however, no bat lyssavirus has been isolated in Asia except for Aravan and Khujand virus in Central Asia. All Chinese lyssavirus isolates in previous reports have been of species rabies virus, mainly from dogs. Following at least two recent bat-associated human rabies-like cases in northeast China, we have initiated a study of the prevalence of lyssaviruses in bats in Jilin province and their public health implications. A bat lyssavirus has been isolated and its pathogenicity in mice and genomic alignment have been determined.

Results

We report the first isolation of a bat lyssavirus in China, from the brain of a northeastern bat, Murina leucogaster. Its nucleoprotein gene shared 92.4%/98.9% (nucleotide) and 92.2%/98.8% (amino acid) identity with the two known Irkut virus isolates from Russia, and was designated IRKV-THChina12. Following intracranial and intramuscular injection, IRKV-THChina12 produced rabies-like symptoms in adult mice with a short inoculation period and high mortality. Nucleotide sequence analysis showed that IRKV-THChina12 has the same genomic organization as other lyssaviruses and its isolation provides an independent origin for the species IRKV.

Conclusions

We have identified the existence of a bat lyssavirus in a common Chinese bat species. Its high pathogenicity in adult mice suggests that public warnings and medical education regarding bat bites in China should be increased, and that surveillance be extended to provide a better understanding of Irkut virus ecology and its significance for public health.

Author Summary

The Lyssavirus genus presently comprises 12 species and two unapproved species with different antigenic characteristics. Rabies virus is detectable worldwide; Lagos bat virus, Mokola virus, Duvenhage virus, Shimoni bat virus, and Ikoma lyssavirus circulate in Africa; European bat lyssavirus types 1 and 2, Irkut virus, West Caucasian bat virus, and Bokeloh bat lyssavirus are found in Europe; and Australian bat lyssavirus has been isolated in Australia. Only Aravan and Khujand viruses have been identified in central Asia. Bats are recognized as the most important reservoirs of lyssaviruses. In China, all lyssavirus isolates in previous reports have been rabies virus, mainly from dogs; none has been from bats. Recently, however, at least two bat-associated human rabies or rabies-like cases have been reported in northeast China. Therefore, we conducted a search for bat lyssaviruses in Jilin province, close to where the first bat-associated human rabies case was recorded. We isolated a bat lyssavirus, identified as an Irkut virus isolate with high pathogenicity in experimental mice. Our findings suggest that public warnings and medical education regarding bat bites in China should be increased, and that surveillance should be extended to provide a better understanding of Irkut virus ecology and its significance for public health.

We conducted a survey in Cambodia in 2000 on henipavirus infection among several bat species, including flying foxes, and persons exposed to these animals. Among 1,072 bat serum samples tested by enzyme-linked immunosorbent assay, antibodies reactive to Nipah virus (NiV) antigen were detected only in Pteropus lylei species; Cynopterus sphinx, Hipposideros larvatus, Scotophilus kuhlii, Chaerephon plicata, Taphozous melanopogon, and T. theobaldi species were negative. Seroneutralization applied on a subset of 156 serum samples confirmed these results. None of the 8 human serum samples was NiV seropositive with the seroneutralization test. One virus isolate exhibiting cytopathic effect with syncytia was obtained from 769 urine samples collected at roosts of P. lylei specimens. Partial molecular characterization of this isolate demonstrated that it was closely related to NiV. These results strengthen the hypothesis that flying foxes could be the natural host of NiV. Surveillance of human cases should be implemented.

Background

Rabies is a fatal encephalitis caused by lyssaviruses. Evidence of lyssavirus circulation has recently emerged in Southeast Asian bats. A cross-sectional study was conducted in Thailand to assess rabies-related knowledge and practices among persons regularly exposed to bats and bat habitats. The objectives were to identify deficiencies in rabies awareness, describe the occurrence of bat exposures, and explore factors associated with transdermal bat exposures.

Methods

A survey was administered to a convenience sample of adult guano miners, bat hunters, game wardens, and residents/personnel at Buddhist temples where mass bat roosting occurs. The questionnaire elicited information on demographics, experience with bat exposures, and rabies knowledge. Participants were also asked to describe actions they would take in response to a bat bite as well as actions for a bite from a potentially rabid animal. Bivariate analysis was used to compare responses between groups and multivariable logistic regression was used to explore factors independently associated with being bitten or scratched by a bat.

Findings

Of 106 people interviewed, 11 (10%) identified bats as a potential source of rabies. A history of a bat bite or scratch was reported by 29 (27%), and 38 (36%) stated either that they would do nothing or that they did not know what they would do in response to a bat bite. Guano miners were less likely than other groups to indicate animal bites as a mechanism of rabies transmission (68% vs. 90%, p = 0.03) and were less likely to say they would respond appropriately to a bat bite or scratch (61% vs. 27%, p = 0.003). Guano mining, bat hunting, and being in a bat cave or roost area more than 5 times a year were associated with history of a bat bite or scratch.

Conclusions

These findings indicate the need for educational outreach to raise awareness of bat rabies, promote exposure prevention, and ensure appropriate health-seeking behaviors for bat-inflicted wounds, particularly among at-risk groups in Thailand.

Author Summary

Rabies is a fatal encephalitis caused by lyssaviruses. Evidence of lyssavirus circulation has recently emerged in Southeast Asian bats. We surveyed persons regularly exposed to bats and bat habitats in Thailand to assess rabies‐related knowledge and practices. Targeted groups included guano miners, bat hunters, game wardens, and residents/personnel at Buddhist temples where mass bat roosting occurs. Of the 106 people interviewed, 11 (10%) identified bats as a source of rabies. History of a bat bite/scratch was reported by 29 (27%), and 38 (36%) expressed either that they would do nothing or that they did not know what they would do in response to a bat bite. Guano miners were less likely than other groups to indicate animal bites as a mechanism of transmission (68% vs. 90%, p=0.03) and were less likely to say they would respond appropriately to a bat bite or scratch (61% vs. 27%, p=0.003). These findings indicate a need for educational outreach in Thailand to raise awareness of bat rabies, promote exposure prevention, and ensure health‐seeking behaviors for bat‐inflicted wounds, particularly among at‐risk groups.

The Aravan virus was isolated from a Lesser Mouse-eared Bat (Myotis blythi) in the Osh region of Kyrghyzstan, central Asia, in 1991. We determined the complete sequence of the nucleoprotein (N) gene and compared it with those of 26 representative lyssaviruses obtained from databases. The Aravan virus was distinguished from seven distinct genotypes on the basis of nucleotide and amino acid identity. Phylogenetic analysis based on both nucleotide and amino acid sequences showed that the Aravan virus was more closely related to genotypes 4, 5, and—to a lesser extent—6, which circulates among insectivorus bats in Europe and Africa. The Aravan virus does not belong to any of the seven known genotypes of lyssaviruses, namely, rabies, Lagos bat, Mokola, and Duvenhage viruses and European bat lyssavirus 1, European bat lyssavirus 2, and Australian bat lyssavirus. Based on these data, we propose a new genotype for the Lyssavirus genus.

Active surveillance for lyssaviruses was conducted among populations of bats in the Philippines. The presence of past or current Lyssavirus infection was determined by use of direct fluorescent antibody assays on bat brains and virus neutralization assays on bat sera. Although no bats were found to have active infection with a Lyssavirus, 22 had evidence of neutralizing antibody against the Australian bat lyssavirus (ABLV). Seropositivity was statistically associated with one species of bat, Miniopterus schreibersi. Results from the virus neutralization assays are consistent with the presence in the Philippines of a naturally occurring Lyssavirus related to ABLV.

The traditional assumption that bats cannot synthesize vitamin C (Vc) has been challenged recently. We have previously shown that two Old World bat species (Rousettus leschenaultii and Hipposideros armiger) have functional L-gulonolactone oxidase (GULO), an enzyme that catalyzes the last step of Vc biosynthesis de novo. Given the uncertainties surrounding when and how bats lost GULO function, exploration of gene evolutionary patterns is needed. We therefore sequenced GULO genes from 16 bat species in 5 families, aiming to establish their evolutionary histories. In five cases we identified pseudogenes for the first time, including two cases in the genus Pteropus (P. pumilus and P. conspicillatus) and three in family Hipposideridae (Coelops frithi, Hipposideros speoris, and H. bicolor). Evolutionary analysis shows that the Pteropus clade has the highest ω ratio and has been subjected to relaxed selection for less than 3 million years. Purifying selection acting on the pseudogenized GULO genes of roundleaf bats (family Hipposideridae) suggests they have lost the ability to synthesize Vc recently. Limited mutations in the reconstructed GULO sequence of the ancestor of all bats contrasts with the many mutations in the ancestral sequence of recently emerged Pteropus bats. We identified at least five mutational steps that were then related to clade origination times. Together, our results suggest that bats lost the ability to biosynthesize vitamin C recently by exhibiting stepwise mutation patterns during GULO evolution that can ultimately lead to pseudogenization.

Brain analysis cannot be used for the investigation of active lyssavirus infection in healthy bats because most bat species are protected by conservation directives. Consequently, serology remains the only tool for performing virological studies on natural bat populations; however, the presence of antibodies merely reflects past exposure to the virus and is not a valid marker of active infection. This work describes a new nested reverse transcription (RT)-PCR technique specifically designed for the detection of the European bat virus 1 on oropharyngeal swabs obtained from bats but also able to amplify RNA from the remaining rabies-related lyssaviruses in brain samples. The technique was successfully used for surveillance of a serotine bat (Eptesicus serotinus) colony involved in a case of human exposure, in which 15 out of 71 oropharyngeal swabs were positive. Lyssavirus infection was detected on 13 oropharyngeal swabs but in only 5 brains out of the 34 animals from which simultaneous brain and oropharyngeal samples had been taken. The lyssavirus involved could be rapidly identified by automatic sequencing of the RT-PCR products obtained from 14 brains and three bat oropharyngeal swabs. In conclusion, RT-PCR using oropharyngeal swabs will permit screening of wild bat populations for active lyssavirus infection, for research or epidemiological purposes, in line not only with conservation policies but also in a more efficient manner than classical detection techniques used on the brain.

In Cambodia, 1,303 bats of 16 species were tested for lyssavirus. No lyssavirus nucleocapsid was detected in 1,283 brains tested by immunofluorescence assay. Antibodies against lyssaviruses were detected by enzyme-linked immunosorbent assay in 144 (14.7%) of 981 serum samples. Thirty of 187 serum samples contained neutralizing antibodies against different lyssaviruses.

Background. Rabies virus (RABV) has circulated in Madagascar at least since the 19th century. Objectives. To assess the circulation of lyssavirus in the island from 2005 to 2010. Materials and Methods. Animal (including bats) and human samples were tested for RABV and other lyssavirus using antigen, ribonucleic acid (RNA), and antibodies detection and virus isolation. Results. Half of the 437 domestic or tame wild terrestrial mammal brains tested were found RABV antigen positive, including 54% of the 341 dogs tested. This percentage ranged from 26% to 75% across the period. Nine of the 10 suspected human cases tested were laboratory confirmed. RABV circulation was confirmed in 34 of the 38 districts sampled. No lyssavirus RNA was detected in 1983 bats specimens. Nevertheless, antibodies against Lagos bat virus were detected in the sera of 12 among 50 Eidolon dupreanum specimens sampled. Conclusion. More than a century after the introduction of the vaccine, rabies still remains endemic in Madagascar.

Daubenton bats may roost infrequently in human dwellings, so risk for human contact is low.

We report the first seroprevalence study of the occurrence of specific antibodies to European bat lyssavirus type 2 (EBLV-2) in Daubenton's bats. Bats were captured from 19 sites across eastern and southern Scotland. Samples from 198 Daubenton's bats, 20 Natterer's bats, and 6 Pipistrelle's bats were tested for EBLV-2. Blood samples (N = 94) were subjected to a modified fluorescent antibody virus neutralization test to determine antibody titer. From 0.05% to 3.8% (95% confidence interval) of Daubenton's bats were seropositive. Antibodies to EBLV-2 were not detected in the 2 other species tested. Mouth swabs (N = 218) were obtained, and RNA was extracted for a reverse transcription–polymerase chain reaction (RT-PCR). The RT-PCR included pan lyssavirus-primers (N gene) and internal PCR control primers for ribosomal RNA. EBLV-2 RNA was not detected in any of the saliva samples tested, and live virus was not detected in virus isolation tests.

Ebolaviruses (EBOV) (family Filoviridae) cause viral hemorrhagic fevers in humans and non-human primates when they spill over from their wildlife reservoir hosts with case fatality rates of up to 90%. Fruit bats may act as reservoirs of the Filoviridae. The migratory fruit bat, Eidolon helvum, is common across sub-Saharan Africa and lives in large colonies, often situated in cities. We screened sera from 262 E. helvum using indirect fluorescent tests for antibodies against EBOV subtype Zaire. We detected a seropositive bat from Accra, Ghana, and confirmed this using western blot analysis. The bat was also seropositive for Lagos bat virus, a Lyssavirus, by virus neutralization test. The bat was fitted with a radio transmitter and was last detected in Accra 13 months after release post-sampling, demonstrating long-term survival. Antibodies to filoviruses have not been previously demonstrated in E. helvum. Radio-telemetry data demonstrates long-term survival of an individual bat following exposure to viruses of families that can be highly pathogenic to other mammal species. Because E. helvum typically lives in large urban colonies and is a source of bushmeat in some regions, further studies should determine if this species forms a reservoir for EBOV from which spillover infections into the human population may occur.

Rabies is a fatal zoonosis caused by a nonsegmented negative-strand RNA virus, namely, rabies virus (RABV). Apart from RABV, at least 10 additional species are known as rabies-related lyssaviruses (RRVs), and some of them are responsible for occasional spillovers into humans. More lyssaviruses have also been detected recently in different bat ecosystems, thanks to the application of molecular diagnostic methods. Due to the variety of the members of the genus Lyssavirus, there is the necessity to develop a reliable molecular assay for rabies diagnosis able to detect and differentiate among the existing rabies and rabies-related viruses. In the present study, a pyrosequencing protocol targeting the 3′ terminus of the nucleoprotein (N) gene was applied for the rapid characterization of lyssaviruses. Correct identification of species was achieved for each sample tested. Results from the pyrosequencing assay were also confirmed by those obtained using the Sanger sequencing method. A pan-lyssavirus one-step reverse transcription (RT)-PCR was developed within the framework of the pyrosequencing procedure. The sensitivity (Se) of the one-step RT-PCR assay was determined by using in vitro-transcribed RNA and serial dilutions of titrated viruses. The assay demonstrated high analytical and relative specificity (Sp) (98.94%) and sensitivity (99.71%). To date, this is the first case in which pyrosequencing has been applied for lyssavirus identification using a cheaper diagnostic approach than the one for all the other protocols for rapid typing that we are acquainted with. Results from this study indicate that this procedure is suitable for lyssavirus detection in samples of both human and animal origin.

The predominant role of Eptesicus serotinus in the epizootic of bat rabies in Europe was further outlined by the first isolation of the rabies virus from this species in France. The distribution of the virus was studied in naturally infected E. serotinus bats at the time of death and suggested that the papillae of the tongue and the respiratory mucosa may play a role in virus production and excretion. The analysis of 501 French rabies virus isolates from various animal species by antinucleocapsid monoclonal antibodies indicated that transmission of the disease from bats to terrestrial animals is unlikely. The antigenic profile of two isolates from French bats corresponded to that of European bat lyssavirus type 1 (EBL1). Comparisons of 12 different isolates from bats with antinucleocapsid and antiglycoprotein monoclonal antibodies and by direct sequencing of the polymerase chain reaction amplification product of the N gene indicated that EBL1, EBL2, Duvenhage virus (serotype 4 of lyssavirus), and the European fox rabies virus (serotype 1) are phylogenetically distant. They formed four tight genetic clusters named genotypes. EBL1 was shown to be antigenically and genetically more closely related to Duvenhage virus than to EBL2. We propose that EBL1 and EBL2 constitute two distinct genotypes which further serologic characterization will probably classify as new serotypes. We also report a simple method for the rapid characterization of EBL based on the digestion of the polymerase chain reaction product of the N gene by three restriction endonucleases.

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Serum specimens from fruit bats were obtained at restaurants in Cambodia. We detected antibodies cross-reactive to Nipah virus by enzyme immunoassay in 11 (11.5%) of 96 Lyle’s flying foxes (Pteropus lylei). Our study suggests that viruses closely related to Nipah or Hendra viruses are more widespread in Southeast Asia than previously documented.

Bats are reservoirs for emerging zoonotic viruses that can have a profound impact on human and animal health, including lyssaviruses, filoviruses, paramyxoviruses, and severe acute respiratory syndrome coronaviruses (SARS-CoVs). In the course of a project focused on pathogen discovery in contexts where human-bat contact might facilitate more efficient interspecies transmission of viruses, we surveyed gastrointestinal tissue obtained from bats collected in caves in Nigeria that are frequented by humans. Coronavirus consensus PCR and unbiased high-throughput pyrosequencing revealed the presence of coronavirus sequences related to those of SARS-CoV in a Commerson’s leaf-nosed bat (Hipposideros commersoni). Additional genomic sequencing indicated that this virus, unlike subgroup 2b CoVs, which includes SARS-CoV, is unique, comprising three overlapping open reading frames between the M and N genes and two conserved stem-loop II motifs. Phylogenetic analyses in conjunction with these features suggest that this virus represents a new subgroup within group 2 CoVs.

IMPORTANCE

Bats (order Chiroptera, suborders Megachiroptera and Microchiroptera) are reservoirs for a wide range of viruses that cause diseases in humans and livestock, including the severe acute respiratory syndrome coronavirus (SARS-CoV), responsible for the global SARS outbreak in 2003. The diversity of viruses harbored by bats is only just beginning to be understood because of expanded wildlife surveillance and the development and application of new tools for pathogen discovery. This paper describes a new coronavirus, one with a distinctive genomic organization that may provide insights into coronavirus evolution and biology.

Abstract

Previous studies have investigated rabies virus (RABV) epizootiology in Brazilian free-tailed bats (Tadarida brasiliensis) in natural cave roosts. However, little is known about geographic variation in RABV exposure, or if the use of man-made roosts by this species affects enzootic RABV infection dynamics within colonies. We sampled rabies viral neutralizing antibodies in bats at three bridge and three cave roosts at multiple time points during the reproductive season to investigate temporal and roost variation in RABV exposure. We report seropositive bats in all age and sex classes with minimal geographic variation in RABV seroprevalence among Brazilian free-tailed bat colonies in south-central Texas. While roost type was not a significant predictor of RABV seroprevalence, it was significantly associated with seasonal fluctuations, suggesting patterns of exposure that differ between roosts. Temporal patterns suggest increased RABV seroprevalence after parturition in cave colonies, potentially related to an influx of susceptible young, in contrast to more uniform seroprevalence in bridge colonies. This study highlights the importance of life history and roost ecology in understanding patterns of RABV seroprevalence in colonies of the Brazilian free-tailed bat.

Six hundred and twenty-eight insectivorous bats originating from seven provinces were submitted to this Institute for rabies diagnosis between August 1, 1963 and December 31, 1967. Brain tissue was examined by the fluorescent antibody technique and the mouse infectivity test was carried out with brain, salivary gland, interscapular adipose tissue and kidney samples. Rabies virus was detected in 44 bats, 29 of which were from Ontario, 12 from British Columbia and three from Manitoba. Most of the positive cases were diagnosed in summer months. Seven species were represented among the specimens found to be rabid; there were 32 big brown bats, three hoary bats, three silver-haired bats, two little brown bats, one eastern pipistrelle, one Keen myotis and one red bat. Another bat which was not identified also proved to be infected with rabies.

In nature, rabies virus (RABV; genus Lyssavirus, family Rhabdoviridae) represents an assemblage of phylogenetic lineages, associated with specific mammalian host species. Although it is generally accepted that RABV evolved originally in bats and further shifted to carnivores, mechanisms of such host shifts are poorly understood, and examples are rarely present in surveillance data. Outbreaks in carnivores caused by a RABV variant, associated with big brown bats, occurred repeatedly during 2001–2009 in the Flagstaff area of Arizona. After each outbreak, extensive control campaigns were undertaken, with no reports of further rabies cases in carnivores for the next several years. However, questions remained whether all outbreaks were caused by a single introduction and further perpetuation of bat RABV in carnivore populations, or each outbreak was caused by an independent introduction of a bat virus. Another question of concern was related to adaptive changes in the RABV genome associated with host shifts. To address these questions, we sequenced and analyzed 66 complete and 20 nearly complete RABV genomes, including those from the Flagstaff area and other similar outbreaks in carnivores, caused by bat RABVs, and representatives of the major RABV lineages circulating in North America and worldwide. Phylogenetic analysis demonstrated that each Flagstaff outbreak was caused by an independent introduction of bat RABV into populations of carnivores. Positive selection analysis confirmed the absence of post-shift changes in RABV genes. In contrast, convergent evolution analysis demonstrated several amino acids in the N, P, G and L proteins, which might be significant for pre-adaptation of bat viruses to cause effective infection in carnivores. The substitution S/T242 in the viral glycoprotein is of particular merit, as a similar substitution was suggested for pathogenicity of Nishigahara RABV strain. Roles of the amino acid changes, detected in our study, require additional investigations, using reverse genetics and other approaches.

Author Summary

Host shifts of the rabies virus (RABV) from bats to carnivores are important for our understanding of viral evolution and emergence, and have significant public health implications, particularly for the areas where “terrestrial” rabies has been eliminated. In this study we addressed several rabies outbreaks in carnivores that occurred in the Flagstaff area of Arizona during 2001–2009, and caused by the RABV variant associated with big brown bats (Eptesicus fuscus). Based on phylogenetic analysis we demonstrated that each outbreak resulted from a separate introduction of bat RABV into populations of carnivores. No post-shift changes in viral genomes were detected under the positive selection analysis. Trying to answer the question why certain bat RABV variants are capable for host shifts to carnivores and other variants are not, we developed a convergent evolution analysis, and implemented it for multiple RABV lineages circulating worldwide. This analysis identified several amino acids in RABV proteins which may facilitate host shifts from bats to carnivores. Precise roles of these amino acids require additional investigations, using reverse genetics and animal experimentation. In general, our approach and the results obtained can be used for prediction of host shifts and emergence of other zoonotic pathogens.

A virus isolated from a Natterer’s bat (Myotis nattererii) in Germany was differentiated from other lyssaviruses on the basis of the reaction pattern of a panel of monoclonal antibodies. Phylogenetic analysis supported the assumption that the isolated virus, Bokeloh bat lyssavirus, may represent a new member of the genus Lyssavirus.

Specimens were obtained from the 3 Malagasy fruit bats, Pteropus rufus, Eidolon dupreanum, and Rousettus madagascariensis. Antibodies against Nipah, Hendra, and Tioman viruses were detected by immunoassay in 23 and by serum neutralization tests in 3 of 427 serum samples, which suggests that related viruses have circulated in Madagascar.

Bat rabies and its transmission to humans and other species in Mexico were investigated. Eighty-nine samples obtained from rabid livestock, cats, dogs, and humans in Mexico were studied by antigenic typing and partial sequence analysis. Samples were further compared with enzootic rabies associated with different species of bats in the Americas. Patterns of nucleotide variation allowed the definition of at least 20 monophyletic clusters associated with 9 or more different bat species. Several lineages associated with distinctive antigenic patterns were found in rabies viruses related to rabies in vampire bats in Mexico. Vampire bat rabies virus lineages associated with antigenic variant 3 are widely spread from Mexico to South America, suggesting these lineages as the most likely ancestors of vampire bat rabies and the ones that have been moved by vampire bat populations throughout the Americas. Rabies viruses related to Lasiurus cinereus, Histiotus montanus, and some other not yet identified species of the genus Lasiurus were found circulating in Mexico. Long-range dissemination patterns of rabies are not necessarily associated with migratory bat species, as in the case of rabies in Desmodus rotundus and Histiotus montanus. Human rabies was associated with vampire bat transmission in most cases, and in one case, rabies transmission from free-tailed bats was inferred. The occurrence of rabies spillover from bats to domestic animals was also demonstrated. Genetic typing of rabies viruses allowed us to distinguish trends of disease dissemination and to address, in a preliminary fashion, aspects of the complex evolution of rabies viruses in different host-reservoir species.

Hipposideros spp. bats harbor a coronavirus that shares common ancestry with human viruses.

We tested 12 bat species in Ghana for coronavirus (CoV) RNA. The virus prevalence in insectivorous bats (n = 123) was 9.76%. CoV was not detected in 212 fecal samples from Eidolon helvum fruit bats. Leaf-nosed bats pertaining to Hipposideros ruber by morphology had group 1 and group 2 CoVs. Virus concentrations were <45,000 copies/100 mg of bat feces. The diversified group 1 CoV shared a common ancestor with the human common cold virus hCoV-229E but not with hCoV-NL63, disputing hypotheses of common human descent. The most recent common ancestor of hCoV-229E and GhanaBt-CoVGrp1 existed in ≈1686–1800 ad. The GhanaBt-CoVGrp2 shared an old ancestor (≈2,400 years) with the severe acute respiratory syndrome–like group of CoV.

Background

Differences among species and among years in reproductive seasonality (the tendency for clusters of events to fall at approximately the same point in each year) and synchrony (amount of clustering of events within a year) have been intensively studied in bats, but are difficult to assess. Here, we use randomization methods with circular statistics to test for synchrony and seasonality of reproduction in three species of nectarivorous megachiropteran bats on Negros Island in the central Philippines.

Results

In Rousettus amplexicaudatus, estimated dates of birth were both highly synchronous and highly seasonal. In Macroglossus minimus, estimated births were seasonal and significantly clustered within years, but within each year births occurred over a broad period, indicating a low level of synchrony. In Eonycteris spelaea, estimated births were also seasonal and had statistically significant synchrony, with birth periods within years intermediate in synchrony between R. amplexicaudatus and M. minimus. All three species had a similar seasonal pattern, with two birth periods in each year, centered on March or April and August or September. In one species, R. amplexicaudatus, primigravid females (in their first pregnancy) produced their young in June and July, a birth period significantly different in timing from the two birth periods of older adult females. This more conservative pattern of young females may allow higher survival of parents and offspring at cost of a lost reproductive opportunity. There was weak evidence that in some years primigravid females of M. minimus might differ in timing from older adults. There were few significant differences in reproductive timing among different years, and those differences were generally less than two weeks, even during a severe drought in the severe el Niño of 1983.

Conclusion

The results suggest that these species follow an obligately seasonal pattern of reproductive timing with very little phenotypic plasticity. The resampling methods were sensitive to differences in timing of under two weeks, in some cases, suggesting that these are useful methods for analyses of seasonality in wild populations of bats.