We screened 217 bats of at least 20 species from 17 locations in Kenya during July and August of 2006 for the presence of adenovirus, rhabdovirus, and paramyxovirus nucleic acids using generic reverse transcription polymerase chain reaction (RT-PCR) and PCR assays. Of 217 bat fecal swabs examined, 4 bats were adenovirus DNA-positive, 11 bats were paramyxovirus RNA-positive, and 2 bats were rhabdovirus RNA-positive. Three bats were coinfected by two different viruses. By sequence comparison and phylogenetic analysis, the Kenya bat paramyxoviruses and rhabdoviruses from this study may represent novel viral lineages within their respective families; the Kenya bat adenoviruses could not be confirmed as novel, because the same region sequences from other known bat adenovirus genomes for comparison were lacking. Our study adds to previous evidence that bats carry diverse, potentially zoonotic viruses and may be coinfected with more than one virus.
Eight years after emigrating from Brazil, an otherwise healthy man developed rabies. An exposure prior to immigration was reported. Genetic analysis revealed a canine rabies virus variant found only in the patient’s home country, and the patient had not traveled internationally since immigrating to the United States. We describe how epidemiological, phylogenetic, and viral sequencing data provided confirmation that rabies encephalomyelitis may present after a long, multiyear incubation period, a consideration that previously has been hypothesized without the ability to exclude a more recent exposure. Accordingly, rabies should be considered in the diagnosis of any acute encephalitis, myelitis, or encephalomyelitis.
In 2009, a novel lyssavirus (subsequently named Ikoma lyssavirus, IKOV) was detected in the brain of an African civet (Civettictis civetta) with clinical rabies in the Serengeti National Park of Tanzania. The degree of nucleotide divergence between the genome of IKOV and those of other lyssaviruses predicted antigenic distinction from, and lack of protection provided by, available rabies vaccines. In addition, the index case was considered likely to be an incidental spillover event, and therefore the true reservoir of IKOV remained to be identified. The advent of sensitive molecular techniques has led to a rapid increase in the discovery of novel viruses. Detecting viral sequence alone, however, only allows for prediction of phenotypic characteristics and not their measurement. In the present study we describe the in vitro and in vivo characterization of IKOV, demonstrating that it is (1) pathogenic by peripheral inoculation in an animal model, (2) antigenically distinct from current rabies vaccine strains and (3) poorly neutralized by sera from humans and animals immunized against rabies. In a laboratory mouse model, no protection was elicited by a licensed rabies vaccine. We also investigated the role of bats as reservoirs of IKOV. We found no evidence for infection among 483 individuals of at least 13 bat species sampled across sites in the Serengeti and Southern Kenya.
The recent discovery of genetically distinct hantaviruses in multiple species of shrews and moles prompted a further exploration of their host diversification by analyzing frozen, ethanol-fixed and RNAlater®-preserved archival tissues and fecal samples from 533 bats (representing seven families, 28 genera and 53 species in the order Chiroptera), captured in Asia, Africa and the Americas in 1981–2012, using RT-PCR. Hantavirus RNA was detected in Pomona roundleaf bats (Hipposideros pomona) (family Hipposideridae), captured in Vietnam in 1997 and 1999, and in banana pipistrelles (Neoromicia nanus) (family Vespertilionidae), captured in Côte d’Ivoire in 2011. Phylogenetic analysis, based on the full-length S- and partial M- and L-segment sequences using maximum likelihood and Bayesian methods, demonstrated that the newfound hantaviruses formed highly divergent lineages, comprising other recently recognized bat-borne hantaviruses in Sierra Leone and China. The detection of bat-associated hantaviruses opens a new era in hantavirology and provides insights into their evolutionary origins.
hantavirus; Chiroptera; evolution
Eidolon helvum; Mokola bat virus; Shimoni bat virus; Lagos bat virus; Chaerophon pumila
The south-central skunk rabies virus (SCSK) is the most broadly distributed terrestrial viral lineage in North America. Skunk rabies has not been efficiently targeted by oral vaccination campaigns and represents a natural system of pathogen invasion, yielding insights to rabies emergence. In the present study we reconstructed spatiotemporal spread of SCSK in the whole territory of its circulation using a combination of Bayesian methods. The analysis based on 241 glycoprotein gene sequences demonstrated that SCSK is much more divergent phylogenetically than was appreciated previously. According to our analyses the SCSK originated in the territory of Texas ~170 years ago, and spread geographically during the following decades. The wavefront velocity in the northward direction was significantly greater than in the eastward and westward directions. Rivers (except the Mississippi River and Rio Grande River) did not constitute significant barriers for epizootic spread, in contrast to deserts and mountains. The mean dispersal rate of skunk rabies was lower than that of the raccoon and fox rabies. Viral lineages circulate in their areas with limited evidence of geographic spread during decades. However, spatiotemporal reconstruction shows that after a long period of stability the dispersal rate and wavefront velocity of SCSK are increasing. Our results indicate that there is a need to develop control measures for SCSK, and suggest how such measure can be implemented most efficiently. Our approach can be extrapolated to other rabies reservoirs and used as a tool for investigation of epizootic patterns and planning interventions towards disease elimination.
Lyssaviruses (family Rhabdoviridae) constitute one of the most important groups of viral zoonoses globally. All lyssaviruses cause the disease rabies, an acute progressive encephalitis for which, once symptoms occur, there is no effective cure. Currently available vaccines are highly protective against the predominantly circulating lyssavirus species. Using next-generation sequencing technologies, we have obtained the whole-genome sequence for a novel lyssavirus, Ikoma lyssavirus (IKOV), isolated from an African civet in Tanzania displaying clinical signs of rabies. Genetically, this virus is the most divergent within the genus Lyssavirus. Characterization of the genome will help to improve our understanding of lyssavirus diversity and enable investigation into vaccine-induced immunity and protection.
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.
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.
The significance of bats as sources of emerging infectious diseases has been increasingly appreciated, and new data have been accumulated rapidly during recent years. For some emerging pathogens the bat origin has been confirmed (such as lyssaviruses, henipaviruses, coronaviruses), for other it has been suggested (filoviruses). Several recently identified viruses remain to be ‘orphan’ but have a potential for further emergence (such as Tioman, Menangle, and Pulau viruses). In the present review we summarize information on major bat-associated emerging infections and discuss specific characteristics of bats as carriers of pathogens (from evolutionary, ecological, and immunological positions). We also discuss drivers and forces of an infectious disease emergence and describe various existing and potential approaches for control and prevention of such infections at individual, populational, and societal levels.
bats; Chiroptera; emerging infectious disease; rabies; lyssavirus; coronavirus; filovirus; henipavirus; prevention; control
TOC summary: Bats may be reservoirs of zoonotic viruses that threaten human health.
Bats are known reservoirs of viral zoonoses. We report genetic characterization of a bat rotavirus (Bat/KE4852/07) detected in the feces of a straw-colored fruit bat (Eidolon helvum). Six bat rotavirus genes (viral protein [VP] 2, VP6, VP7, nonstructural protein [NSP] 2, NSP3, and NSP5) shared ancestry with other mammalian rotaviruses but were distantly related. The VP4 gene was nearly identical to that of human P rotavirus strains, and the NSP4 gene was closely related to those of previously described mammalian rotaviruses, including human strains. Analysis of partial sequence of the VP1 gene indicated that it was distinct from cognate genes of other rotaviruses. No sequences were obtained for the VP3 and NSP1 genes of the bat rotavirus. This rotavirus was designated G25-P-I15-R8(provisional)-C8-Mx-Ax-N8-T11-E2-H10. Results suggest that several reassortment events have occurred between human, animal, and bat rotaviruses. Several additional rotavirus strains were detected in bats.
Straw-colored fruit bat; Eidolon helvum; rotavirus; viruses; reassortment; heterologous genome segments; podcast; zoonoses; research
We report the presence and diversity of Bartonella spp. in bats of 13 insectivorous and frugivorous species collected from various locations across Kenya. Bartonella isolates were obtained from 23 Eidolon helvum, 22 Rousettus aegyptiacus, 4 Coleura afra, 7 Triaenops persicus, 1 Hipposideros commersoni, and 49 Miniopterus spp. bats. Sequence analysis of the citrate synthase gene from the obtained isolates showed a wide assortment of Bartonella strains. Phylogenetically, isolates clustered in specific host bat species. All isolates from R. aegyptiacus, C. afra, and T. persicus bats clustered in separate monophyletic groups. In contrast, E. helvum and Miniopterus spp. bats harbored strains that clustered in several groups. Further investigation is needed to determine whether these agents are responsible for human illnesses in the region.
Bacteria; Bartonella; bats; zoonoses; Kenya; research
Lake Victoria Marburgvirus; Marburg virus; bats; Egyptian fruit bat; Rousettus aegyptiacus; zoonosis; Kenya; filovirus; viruses; letter
Diverse coronaviruses have been identified in bats from several continents but not from Africa. We identified group 1 and 2 coronaviruses in bats in Kenya, including SARS-related coronaviruses. The sequence diversity suggests that bats are well-established reservoirs for and likely sources of coronaviruses for many species, including humans.
Coronavirus; SARS; bat viruses; pathogen discovery; emerging viral infections; dispatch
The prevalence of neutralizing antibody against West Caucasian bat virus (WCBV) in Miniopterus bats collected in Kenya ranged from 17% to 26%. Seropositive bats were detected in 4 of 5 locations sampled across the country. These findings provide evidence that WCBV, originally isolated in Europe, may emerge in other continents.
West Caucasian bat virus, lyssavirus; bats, Miniopterus, seroprevalence, dispatch
During lyssavirus surveillance, 1,221 bats of at least 30 species were collected from 25 locations in Kenya. One isolate of Lagos bat virus (LBV) was obtained from a dead Eidolon helvum fruit bat. The virus was most similar phylogenetically to LBV isolates from Senegal (1985) and from France (imported from Togo or Egypt; 1999), sharing with these viruses 100% nucleoprotein identity and 99.8 to 100% glycoprotein identity. This genome conservancy across space and time suggests that LBV is well adapted to its natural host species and that populations of reservoir hosts in eastern and western Africa have sufficient interactions to share pathogens. High virus concentrations, in addition to being detected in the brain, were detected in the salivary glands and tongue and in an oral swab, suggesting that LBV is transmitted in the saliva. In other extraneural organs, the virus was generally associated with innervations and ganglia. The presence of infectious virus in the reproductive tract and in a vaginal swab implies an alternative opportunity for transmission. The isolate was pathogenic for laboratory mice by the intracerebral and intramuscular routes. Serologic screening demonstrated the presence of LBV-neutralizing antibodies in E. helvum and Rousettus aegyptiacus fruit bats. In different colonies the seroprevalence ranged from 40 to 67% and 29 to 46% for E. helvum and R. aegyptiacus, respectively. Nested reverse transcription-PCR did not reveal the presence of viral RNA in oral swabs of bats in the absence of brain infection. Several large bat roosts were identified in areas of dense human populations, raising public health concerns for the potential of lyssavirus infection.
Canine rabies is a neglected disease causing 55,000 human deaths worldwide per year, and 99% of all cases are transmitted by dog bites. In N'Djaména, the capital of Chad, rabies is endemic with an incidence of 1.71/1,000 dogs (95% C.I. 1.45–1.98). The gold standard of rabies diagnosis is the direct immunofluorescent antibody (DFA) test, requiring a fluorescent microscope. The Centers for Disease Control and Prevention (CDC, Atlanta, United States of America) developed a histochemical test using low-cost light microscopy, the direct rapid immunohistochemical test (dRIT).
We evaluated the dRIT in the Chadian National Veterinary Laboratory in N'Djaména by testing 35 fresh samples parallel with both the DFA and dRIT. Additional retests (n = 68 in Chad, n = 74 at CDC) by DFA and dRIT of stored samples enhanced the power of the evaluation. All samples were from dogs, cats, and in one case from a bat. The dRIT performed very well compared to DFA. We found a 100% agreement of the dRIT and DFA in fresh samples (n = 35). Results of retesting at CDC and in Chad depended on the condition of samples. When the sample was in good condition (fresh brain tissue), we found simple Cohen's kappa coefficient related to the DFA diagnostic results in fresh tissue of 0.87 (95% C.I. 0.63–1) up to 1. For poor quality samples, the kappa values were between 0.13 (95% C.I. −0.15–0.40) and 0.48 (95% C.I. 0.14–0.82). For samples stored in glycerol, dRIT results were more likely to agree with DFA testing in fresh samples than the DFA retesting.
The dRIT is as reliable a diagnostic method as the gold standard (DFA) for fresh samples. It has an advantage of requiring only light microscopy, which is 10 times less expensive than a fluorescence microscope. Reduced cost suggests high potential for making rabies diagnosis available in other cities and rural areas of Africa for large populations for which a capacity for diagnosis will contribute to rabies control.
A new diagnostic test for rabies in animals was evaluated in N'Djaména, capital of Chad. The test is based on a direct immuno-histochemical detection of rabies virus in brain tissue (dRIT) visible by normal light microscopy. Rabies detection by dRIT light microscopy is 10 times less expensive than fluorescence microscopy required for the current gold standard of rabies diagnosis. The test showed ideal results in fresh samples with 100% agreement with the gold standard and confirms the results of a first study in Tanzania. Thus, it has a significant potential for diagnosing rabies in low-income countries, and under field conditions where rabies diagnosis is unavailable for the moment. This new test opens up a great potential to train technical staff and to establish rabies diagnosis without delay in low-income countries with urban rabies.
One-sentence summary for table of contents: Lagos bat virus from water mongoose showed strong sequence homology with other Lagos bat virus isolates from South Africa.
A genotype 2 lyssavirus, Lagos bat virus (LBV), was isolated from a terrestrial wildlife species (water mongoose) in August 2004 in the Durban area of the KwaZulu-Natal Province of South Africa. The virus isolate was confirmed as LBV by antigenic and genetic characterization, and the mongoose was identified as Atilax paludinosus by mitochondrial cytochrome b sequence analysis. Phylogenetic analysis demonstrated sequence homology with previous LBV isolates from South African bats. Studies performed in mice indicated that the peripheral pathogenicity of LBV had been underestimated in previous studies. Surveillance strategies for LBV in Africa must be improved to better understand the epidemiology of this virus and to make informed decisions on future vaccine strategies because evidence is insufficent that current rabies vaccines provide protection against LBV.
Lagos bat virus; rabies; rabies-related viruses; lyssaviruses; nucleoprotein; Herpestidae; mongoose; Atilax paludinosus; South Africa; cytochrome b; pathogenesis; phylogeny; research
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.
lyssavirus; Khujand virus; Aravan virus; rabies; bat; rhabdovirus; Bangladesh; tropical Asia; antibody; dispatch
Surveillance for lyssaviruses was conducted among bat populations in 8 provinces in Thailand. In 2002 and 2003, a total of 932 bats of 11 species were captured and released after serum collection. Lyssavirus infection was determined by conducting virus neutralization assays on bat serum samples. Of collected samples, 538 were either hemolysed or insufficient in volume, which left 394 suitable for analysis. These samples included the following: Pteropus lylei (n = 335), Eonycteris spelaea (n = 45), Hipposideros armiger (n = 13), and Rousettus leschennaulti (n = 1). No serum samples had evidence of neutralizing antibodies when tested against rabies virus. However, 16 samples had detectable neutralizing antibodies against Aravan virus, Khujand virus, Irkut virus, or Australian bat lyssavirus; all were specifically associated with fruit bats P. lylei (n = 15) and E. spelaea (n = 1). These results are consistent with the presence of naturally occurring viruses related to new putative lyssavirus genotypes.
Lyssavirus; rabies; RNA; bat; chiroptera; zoonosis; animals; fluorescent antibody technique; direct/veterinary; Thailand; research
With rabies emerging as a particular threat to wild canids, we report on a rabies outbreak in a subpopulation of endangered Ethiopian wolves in the Bale Mountains, Ethiopia, in 2003 and 2004. Parenteral vaccination of wolves was used to manage the outbreak.
Canis simensis; endangered species; epidemic; Ethiopian wolves; rabies; wildlife diseases; dispatch
Two Nipah virus encephalitis outbreaks in Bangladesh may be associated with person-to-person transmission.
We retrospectively investigated two outbreaks of encephalitis in Meherpur and Naogaon, Bangladesh, which occurred in 2001 and 2003. We collected serum samples from persons who were ill, their household contacts, randomly selected residents, hospital workers, and various animals. Cases were classified as laboratory confirmed or probable. We identified 13 cases (4 confirmed, 9 probable) in Meherpur; 7 were in persons in two households. Patients were more likely than nonpatients to have close contact with other patients or have contact with a sick cow. In Naogaon, we identified 12 cases (4 confirmed, 8 probable); 7 were in persons clustered in 2 households. Two Pteropus bats had antibodies for Nipah virus. Samples from hospital workers were negative for Nipah virus antibodies. These outbreaks, the first since 1999, suggest that transmission may occur through close contact with other patients or from exposure to a common source. Surveillance and enhancement of diagnostic capacity to detect Nipah virus infection are recommended.
Two new rabies-related viruses were discovered in Russia during 2002. Viruses were isolated from bats in Eastern Siberia near Baikal Lake and in the western Caucasus Mountains. After preliminary antigenic and genetic characterization, we found that both viruses should be considered as new putative lyssavirus genotypes.
lyssavirus; rabies; bat; rhabdovirus; infectious disease; Russia; Eurasia; virus; encephalitis
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.
Lyssavirus; genotype; N gene; phylogenetic analysis; bat; central Asia; research