N. Johnson

European bat lyssaviruses: an emerging zoonosis.

In Europe, two bat lyssaviruses referred to as European bat lyssaviruses (EBLVs) types 1 and 2 (genotypes 5 and 6 respectively) which are closely related to classical rabies virus are responsible for an emerging zoonosis. EBLVs are host restricted to bats, and have been known to infect not only their primary hosts but also in rare circumstances, induce spillover infections to terrestrial mammals including domestic livestock, wildlife and man. Although spillover infections have occurred, there has been no evidence that the virus adapted to a new host. Since 1977, four human deaths from EBLVs have been reported. None of them had a record of prophylactic rabies immunization. Only fragmentary data exist about the effectiveness of current vaccines in cross-protection against EBLVs. It is clear that EBLV in bats cannot be eliminated using conventional strategies similar to the control programmes based on vaccine baits used for fox rabies in Europe during the 1980s. Due to the protected status of bats in Europe, our knowledge of EBLV prevalence and epidemiology is limited. It is possible that EBLV is under-reported and that the recorded cases of EBLV represent only a small proportion of the actual number of infected bats. For this reason, any interaction between man and bats in Europe must be considered as a possible exposure. Human exposure through biting incidents, especially unprovoked attacks, should be treated immediately with rabies post-exposure treatment and the bat, where possible, retained for laboratory analysis. Preventative measures include educating all bat handlers of the risks posed by rabies-infected animals and advising them to be immunized. This review provides a brief history of EBLVs, their distribution in host species and the public health risks.

Phylogenetic comparison of the genus Lyssavirus using distal coding sequences of the glycoprotein and nucleoprotein genes

Summary.u2002The phylogenetic relationships between all seven genotypes within the genus Lyssavirus were compared at the nucleotide level utilising two distal regions of the viral genome. The resulting analysis of each region produced similar, although not identical, phylogenetic results, suggesting that the evolutionary pressures on individual proteins within the genome vary. These differences are in part due to the increased variability observed within the glycoprotein sequence over the nucleoprotein sequence. Pair-wise comparison using the glycoprotein partial sequence between different isolates demonstrate that within genotypes, viruses show between 80 and 100% sequence identity, whilst between genotypes, viruses show between 50 and 75% identity. This provides a consistent guide to assigning new viruses to existing genotypes. Alignment of the amino acid sequence for the truncated glycoprotein sequence to the Pasteur Virus vaccine strain show significant residue variation between positions 139 and 170. However, residue variation tends to vary with genotype implying that these changes have not evolved due to immunological pressure from the host but have occurred following the separation of viruses into discrete groups. Comparison of the phylogenetic analysis for this partial region of the glycoprotein suggest that it gives comparable results to studies that have used larger regions of the Lyssavirus genome.

Human rabies due to lyssavirus infection of bat origin.

Rabies is a fatal viral encephalitis and results from infection with viruses belonging to the genus Lyssavirus. Infection usually results from a bite from a dog infected with classical rabies virus. However, a small number of cases result from contact with bats. It is within bats that most lyssavirus variants, referred to as genotypes, are found. The lyssaviruses found in bats have a distinct geographical distribution and are often restricted to specific bat species. Most have been associated with rabies in humans and in some cases spill-over to domestic animals. Many diagnostic techniques are unable to differentiate rabies virus from other genotypes so it is possible that some human and animal cases go unreported. Furthermore, current vaccines have limited efficacy against some genotypes.

Epidemiology of bat rabies in Germany

Summary.Rabies in European bats was first reported in Germany in 1954. In concordance with Denmark and the Netherlands, Germany has reported one of the highest numbers (n = 187) of European bat lyssavirus (EBLV)-positive cases in bats in Europe so far (1954–2005). A combined descriptive epidemiological and phylogenetic analysis on bat rabies and prevailing EBLVs is presented, comprising the past 50 years. So far, only the two lineages of EBLV-1 (genotype 5), a and b, have been detected. Although only 50% of the rabies-positive bats have been identified by species, the Serotine bat (Eptesicus serotinus) is the bat species most frequently infected. Single rabies cases have also been detected in a further five indigenous bat species. There is proven evidence for a substantial bias in the frequency of bat rabies cases in the north of Germany, with an endemic cluster in the northwesternmost low-lying plain areas adjacent to the Netherlands and Denmark. Improvements to bat rabies surveillance and research are discussed.

Detection of mosquito-only flaviviruses in Europe.

The genus Flavivirus, family Flaviviridae, includes a number of important arthropod-transmitted human pathogens such as dengue viruses, West Nile virus, Japanese encephalitis virus and yellow fever virus. In addition, the genus includes flaviviruses without a known vertebrate reservoir, which have been detected only in insects, particularly in mosquitoes, such as cell fusing agent virus, Kamiti River virus, Culex flavivirus, Aedes flavivirus, Quang Binh virus, Nakiwogo virus and Calbertado virus. Reports of the detection of these viruses with no recognized pathogenic role in humans are increasing in mosquitoes collected around the world, particularly in those sampled in entomological surveys targeting pathogenic flaviviruses. The presence of six potential flaviviruses, detected from independent European arbovirus surveys undertaken in the Czech Republic, Italy, Portugal, Spain and the UK between 2007 and 2010, is reported in this work. Whilst the Aedes flaviviruses, detected in Italy from Aedes albopictus mosquitoes, had already been isolated in Japan, the remaining five viruses have not been reported previously: one was detected in Italy, Portugal and Spain from Aedes mosquitoes (particularly from Aedes caspius), one in Portugal and Spain from Culex theileri mosquitoes, one in the Czech Republic and Italy from Aedes vexans, one in the Czech Republic from Aedes vexans and the last in the UK from Aedes cinereus. Phylogenetic analysis confirmed the close relationship of these putative viruses to other insect-only flaviviruses.

Isolation of a European bat lyssavirus type 2 from a Daubenton's bat in the United Kingdom

European bat lyssavirus type 2 (EBLV-2) has been isolated once previously from a bat in the UK in June 1996. In September 2002, a Daubentons bat (Myotis daubentonif) found in Lancashire developed abnormal behaviour, including unprovoked aggression, while it was in captivity. Brain samples from the bat were tested for virus of the Lyssavirus genus, which includes EBLV-2 (genotype 6), and classical rabies virus (genotype 1). A positive fluorescent antibody test confirmed that it was infected with a lyssavirus, and PCR and genomic sequencing identified the virus as an EBLV-2a. Phylogenetic comparisons with all the published sequences from genotype 6 showed that it was closely related to the previous isolate of EBLV-2 in the UK and suggested links to isolates from bats in the Netherlands. The isolation of EBLV-2 from a bat found on the west coast of England provides evidence that this virus may be present within the UK Daubentons bat population at a low prevalence level.

Experimental infection of serotine bats (Eptesicus serotinus) with European bat lyssavirus type 1a.

The serotine bat (Eptesicus serotinus) accounts for the vast majority of bat rabies cases in Europe and is considered the main reservoir for European bat lyssavirus type 1 (EBLV-1, genotype 5). However, so far the disease has not been investigated in its native host under experimental conditions. To assess viral virulence, dissemination and probable means of transmission, captive bats were infected experimentally with an EBLV-1a virus isolated from a naturally infected conspecific from Germany. Twenty-nine wild caught bats were divided into five groups and inoculated by intracranial (i.c.), intramuscular (i.m.) or subcutaneous (s.c.) injection or by intranasal (i.n.) inoculation to mimic the various potential routes of infection. One group of bats was maintained as uninfected controls. Mortality was highest in the i.c.-infected animals, followed by the s.c. and i.m. groups. Incubation periods varied from 7 to 26 days depending on the route of infection. Rabies did not develop in the i.n. group or in the negative-control group. None of the infected bats seroconverted. Viral antigen was detected in more than 50% of the taste buds of an i.c.-infected animal. Shedding of viable virus was measured by virus isolation in cell culture for one bat from the s.c. group at 13 and 14 days post-inoculation, i.e. 7 days before death. In conclusion, it is postulated that s.c. inoculation, in nature caused by bites, may be an efficient way of transmitting EBLV-1 among free-living serotine bats.

Comparative Pathological Study of the Murine Brain after Experimental Infection with Classical Rabies Virus and European Bat Lyssaviruses

European bat lyssaviruses (EBLVs) types 1 (EBLV-1) and 2 (EBLV-2) cause rabies in terrestrial species, but the pathological changes associated with neuroinvasion have yet to be fully elucidated. Swiss OF-1 mice were inoculated peripherally with strain RV61 (classical rabies virus), RV1423 (EBLV-1) or RV1332 (EBLV-2) to compare the nature and extent of histopathological changes produced. Inoculated animals developed varying degrees of non-suppurative encephalitis, and lyssavirus infection was confirmed by the detection of viral antigen. The lesions produced, which included perivascular cuffs and gliosis, were more severe after RV1423 or RV1332 infection than after RV61 infection. Perivascular cuffs were mainly localized to caudal brain regions, irrespective of the infecting strain; after RV1332 infection, however, they were particularly abundant, being composed of large numbers of inflammatory cells. T cells were the predominant lymphocytic component of the inflammatory infiltrate in both the Virchow-Robin space and the brain parenchyma. Viral antigen, which was widespread throughout the brain, was apparently unrelated to the degree of cuffing. The study suggested that there was increased immune activation after inoculation with strain RV1423 or RV1332, particularly the latter, but that this did not affect the final outcome.

European bat lyssaviruses — an ecological enigma

ABSTRACT The first European case of bat rabies was reported in 1954. Since then, more than 800 cases have been confirmed from 13 countries. The causative agents are European Bat Lyssavirus Type-1 (EBLV-1) and Type-2 (EBLV-2). The natural host of EBLV-1 seems to be the serotine bat (Eptesicus serotinus) with more than 95% of all cases reported in this species. Although much fewer EBLV-2 cases have been identified, it seems that Myotis dasycneme and M. daubentonii are likely to be the principal hosts species for this virus. The ecological mechanisms underlying the transmission of EBLV-1 and EBLV-2 are still poorly understood. In order to assess the public health risks associated with EBLVs and the implications for bat conservation, some issues need urgent clarification. In this review some of the most pressing topics will be addressed. Only collaboration between a range of disciplines that include virologists, epidemiologists and bat conservationists will be able to elucidate some of these unanswered questions.

Susceptibility of North American big brown bats (Eptesicus fuscus) to infection with European bat lyssavirus type 1

The aim of this study was to determine the susceptibility of insectivorous bats (using the big brown bat as a model) to infection with European bat lyssavirus type 1a (EBLV-1a), to assess the dynamics of host immune responses and to evaluate the opportunity for horizontal viral transmission within colonies. Two isolates of EBLV-1a, originating from Slovakia (EBLV-1aSK) and Germany (EBLV-1aGE), were tested. Four different routes of inoculation were used with isolate EBLV-1aSK [10(4.8) mouse intracerebral median lethal dose (MICLD(50)) in 50 mul]: intramuscular (i.m.) in the deltoid area or masseter region, per os (p.o.) and intradermal (i.d.) scratches. Isolate EBLV-1aGE (10(3.2) and 10(2.2) MICLD(50) in 20 mul) was inoculated via the intranasal (i.n.), i.m. (low- and high-dose groups, into pectoral muscles); p.o. and intracerebral (i.c.) routes. None of the bats infected by the i.n., p.o. or i.d. route with either virus isolate developed disease during the experiments (91 or 120 days, respectively). Incubation periods were 9-12 days for i.c.-inoculated bats (66 % mortality), 12-33 days for bats inoculated i.m. with the higher dose (23-50 % mortality) and 21-58 days in bats inoculated i.m. with the lower dose of virus (57 % mortality). Virus or viral RNA in bat saliva was detected occasionally, as early as 37 days before death. All i.d.-inoculated and the majority of i.m.-inoculated bats seroconverted within 7-10 days of inoculation. These observations suggest that exposure of bats to varying doses of EBLV-1 from rabid conspecifics via natural (i.d.) routes could lead to an abortive infection and serve as a natural mode of immunization resulting in the presence of virus-neutralizing antibodies in free-ranging bats.