Rabies Virus-Neutralizing Antibodies in Free-Ranging Invasive Wild Boars (Sus scrofa) from Brazil.

Rabies, one of the most lethal global zoonoses, affects all mammals. It remains circulating worldwide in sylvatic cycles through terrestrial and airborne reservoirs, and in Brazil, bats are currently the main reservoirs and source of transmission. Wild boars, an important invasive alien species in Brazil, are a proven food source for hematophagous bats and may participate in the Brazilian sylvatic cycle of rabies. We evaluated the presence of this pathogen in hunted wild boars from the São Paulo state using histopathology, the direct fluorescent antibody test (DFA), viral isolation in cell culture (VICC), the rapid fluorescent focus inhibition test (RFFIT), and quantitative reverse transcription polymerase chain reaction (RT-qPCR). The results of histopathological, DFA, VICC, and RT-qPCR analysis were negative for all samples; seven serum samples tested positive in the RFFIT, and titers ranged from 0.13 IU/mL to 0.5 IU/mL. The presence of rabies virus-neutralizing antibodies in the studied wild boars suggests the circulation of the virus in these animals. Educative actions directed at hunters should include information on the prevention of this important zoonosis.

Detection of rabies virus in cranial cavity lavage of naturally infected bats.

The rabies virus (RABV) has been isolated in several bats species in the world, and among them, hematophagous, frugivorous and insectivorous species. Bats found in Brazil are small, which can lead to situations in which there are limitations in the collection of the central nervous system (CNS) and the amount of material may be insufficient to carry out laboratory diagnostic techniques for rabies. The objective of this work was to evaluate an alternative sample collection for the diagnosis of rabies in bats. A total of 92 bat samples, 82 positives and 10 negatives were selected. The cranial cavity was scraped with the aid of sterile tips and a virus diluent was added to create a suspension. All samples were submitted to Rabies Tissue Culture Infection Test (RTCIT) and reverse transcription polymerase chain reaction (RT-PCR). The diagnostic sensitivity and specificity of the RTCIT and RT-PCR using the cranial cavity lavage were calculated in comparison with the results of the laboratory routine (DFAT and RTCIT) performed with the CNS (considered gold standard). The results of the RTCIT show that the cranial cavity lavage is not an adequate sample for viral isolation, since the diagnostic sensitivity was low (37.8 %) when compared with the tests with the CNS. However, the RT-PCR of the cranial cavity lavage may be a tool to assist in the diagnosis, since it presented a sensitivity of 76.8 %. The results of this study suggest that cranial cavity lavage is an interesting alternative to enable the diagnosis of rabies in bats and increases the possibility of diagnosis contributing to rabies surveillance and control.

Phylogeography of rabies isolated from cattle and horses transmitted by haematophagous bat Desmodus Rotundus in Brazil

Rabies is Public Health problem and is very important in Animal Health too. The illness demands continuous prophylactic care for herbivores with economic interest, such as cattle and horses. The main vector of rabies virus (RABV) for these animals are the hematophagous bats Desmodus rotundus. RABV is a RNA genome with low level of fidelity during replication cycle due to lack of repair of its polymerase. This causes the incorporation of mutations that increase the genotypic variation of the viral population. In the project, the nucleoprotein (N) gene of the RABV isolated mainly from cattle in different cities of State of São Paulo (SP), will be sequenced. In addition, genetic sequences deposited in GenBank will be also used. N is the most conserved gene of RABV for these reason is the most appropriated for phylogeographic studies. Because the RABV display evolutionary and ecological dynamics on the same time scale reliable phylogeographic inferences can be obtained from molecular data. As phylogeography expresses the contemporary pattern of geographic distribution of an organism according to gene genealogies the objective of this project is to determine the dispersion over time and space of the RABV transmitted by D. rotundus in SP. The phylogeography of RABV will be studied by phylogenetic analysis using Bayesian statistics using Monte Carlo methods via Markov Chains (MCMC), available on the Bayesian Evolutionary Analysis Sampling Trees (BEAST) plataform. In this way and after the test of different evolutionary models the data of the phylogenetic trees of substitution and more probable time will be converted into a KML file that allows the visualization of the spatial projection of the diffusion of the genetic lineages in the time and space using Google Earth. In this way, the final results can aid epidemiological surveillance and also strategic planning for the control of rabies.

A rabies virus vampire bat variant shows increased neuroinvasiveness in mice when compared to a carnivore variant

Rabies is one of the most important zoonotic diseases and is caused by several rabies virus (RABV) variants. These variants can exhibit differences in neurovirulence, and few studies have attempted to evaluate the neuroinvasiveness of variants derived from vampire bats and wild carnivores. The aim of this study was to evaluate the neuropathogenesis of infection with two Brazilian RABV street variants (variant 3 and crab-eating fox) in mice. BALB/c mice were inoculated with RABV through the footpad, with the 50% mouse lethal dose (LD50) determined by intracranial inoculation. The morbidity of rabies in mice infected with variant 3 and the crab-eating fox strain was 100% and 50%, respectively, with an incubation period of 7 and 6 days post-inoculation (dpi), respectively. The clinical disease in mice was similar with both strains, and it was characterized initially by weight loss, ruffled fur, hunched posture, and hind limb paralysis progressing to quadriplegia and recumbency at 9 to 12 dpi. Histological lesions within the central nervous system (CNS) characterized by nonsuppurative encephalomyelitis with neuronal degeneration and necrosis were observed in mice infected with variant 3 and those infected with the crab-eating fox variant. However, lesions and the presence of RABV antigen, were more widespread within the CNS of variant-3-infected mice, whereas in crab-eating fox-variant-infected mice, RABV antigens were more restricted to caudal areas of the CNS, such as the spinal cord and brainstem. In conclusion, the results shown here demonstrate that the RABV vampire bat strain (variant 3) has a higher potential for neuroinvasiveness than the carnivore variant. (AU) i

A rabies virus vampire bat variant shows increased neuroinvasiveness in mice when compared to a carnivore variant.

Rabies is one of the most important zoonotic diseases and is caused by several rabies virus (RABV) variants. These variants can exhibit differences in neurovirulence, and few studies have attempted to evaluate the neuroinvasiveness of variants derived from vampire bats and wild carnivores. The aim of this study was to evaluate the neuropathogenesis of infection with two Brazilian RABV street variants (variant 3 and crab-eating fox) in mice. BALB/c mice were inoculated with RABV through the footpad, with the 50% mouse lethal dose (LD50) determined by intracranial inoculation. The morbidity of rabies in mice infected with variant 3 and the crab-eating fox strain was 100% and 50%, respectively, with an incubation period of 7 and 6 days post-inoculation (dpi), respectively. The clinical disease in mice was similar with both strains, and it was characterized initially by weight loss, ruffled fur, hunched posture, and hind limb paralysis progressing to quadriplegia and recumbency at 9 to 12 dpi. Histological lesions within the central nervous system (CNS) characterized by nonsuppurative encephalomyelitis with neuronal degeneration and necrosis were observed in mice infected with variant 3 and those infected with the crab-eating fox variant. However, lesions and the presence of RABV antigen, were more widespread within the CNS of variant-3-infected mice, whereas in crab-eating fox-variant-infected mice, RABV antigens were more restricted to caudal areas of the CNS, such as the spinal cord and brainstem. In conclusion, the results shown here demonstrate that the RABV vampire bat strain (variant 3) has a higher potential for neuroinvasiveness than the carnivore variant.

Diphylla ecaudata y Diaemus youngi, Biología y comportamiento / Diphylla ecaudata and Diaemus youngi, Biology and behavior

En la presente revisión se presentan algunos aspectos de la biología y comportamiento de las especies de quirópteros hematófagos Diphylla ecaudata y Diaemus youngi. Y se analizan las principales características anatómicas que permiten hacer la diferenciación de estas especies con Desmodus rotundus. Es notable la falta de información sobre las dos primeras especies y la necesidad de capacitación para la correcta identificación de las mismas, lo que redundará en un direccionamiento más específico para controlar las poblaciones de Desmodus rotundus.

This review presents some aspects about the biology and behavior of the hematophagous bats Diphylla ecaudata and Diaemus youngi. It is notable the lack of information regarding both species, and the need of training in order to identify them properly from Desmodus rotundus. We analyze the main anatomical features that allow differentiation of these species. This will result in a more specific control of Desmodus rotundus populations.

Human rabies transmitted by vampire bats: antigenic and genetic characterization of rabies virus isolates from the Amazon region (Brazil and Ecuador).

Since 2004, the main transmitter of human rabies in Latin America has been the vampire bat (Desmodus rotundus). Based on the nucleoprotein of the rabies virus (RV), we analyzed antigenic and genetic profiles of isolates from 29 samples taken from humans living in different areas of the Amazon region. Two isolates were from Ecuador and 27 from the Northern and Northeastern regions of Brazil, which were obtained during outbreaks in various municipalities in the states of Pará and Maranhão in the years 2004 and 2005. The partial N gene (nt 104-1477) of the 29 isolates was sequenced, and the sequences were used to build a neighbor-joining tree with the Kimura-2 parameter model. All 29 human RV isolates were identified as belonging to antigenic variant 3 (AgV3) and were genetically grouped into the D. rotundus cluster, which was divided into two subclusters (A and B), subcluster A in turn being divided into four genetic groups (A1, A2, A3 and A4). Genetic and molecular markers characterizing these genetic lineages were also identified. The results of this study show that the isolates belong to the same rabies cycle as that of the vampire bat D. rotundus. However, the division of clusters within the lineage associated with D. rotundus shows that different genetic sublineages of the virus were circulating in the Amazon region during the study period. Our findings suggest that there are phylogeographic differences between isolates obtained over a short period.

Genetic characterization of rabies virus isolated from bovines and equines between 2007 and 2008, in the States of São Paulo and Minas Gerais.

INTRODUCTION: Rabies is an acute disease of the central nervous system and is responsible for the deaths of thousands of humans, wild animals and livestock, particularly cattle, as well as causing major economic losses. This study describes the genetic characterization of rabies virus variants that circulate in Desmodus rotundus populations and are transmitted to herbivores. METHODS: Fifty rabies virus isolates from bovines and equines in the States of São Paulo and Minas Gerais, Brazil, were genetically characterized and compared with sequences retrieved from GenBank. RESULTS: Two clusters (I and II) with mean nucleotide identities of 99.1 and 97.6% were found. The first of these contained nearly all the samples analyzed. Lineages from other Brazilian states grouped in cluster II. CONCLUSIONS: Analysis of the amino acid sequences of the N proteins revealed the existence of genetic markers that may indicate possible variations between geographic regions, although the biologically active regions are conserved within the species over space and time.

Comparative analysis of rabies virus isolates from Brazilian canids and bats based on the G gene and G-L intergenic region.

Rabies virus (RABV) isolates from two species of canids and three species of bats were analyzed by comparing the C-terminal region of the G gene and the G-L intergenic region of the virus genome. Intercluster identities for the genetic sequences of the isolates showed both regions to be poorly conserved. Phylogenetic trees were generated by the neighbor-joining and maximum parsimony methods, and the results were found to agree between the two methods for both regions. Putative amino acid sequences obtained from the G gene were also analyzed, and genetic markers were identified. Our results suggest that different genetic lineages of RABV are adapted to different animal species in Brazil.

Genetic characterization of rabies virus isolated from bovines and equines between 2007 and 2008, in the States of São Paulo and Minas Gerais / Caracterização genética de vírus da raiva isolado de bovinos e equinos entre 2007 e 2008, nos Estados de São Paulo e Minas Gerais

INTRODUCTION: Rabies is an acute disease of the central nervous system and is responsible for the deaths of thousands of humans, wild animals and livestock, particularly cattle, as well as causing major economic losses. This study describes the genetic characterization of rabies virus variants that circulate in Desmodus rotundus populations and are transmitted to herbivores. METHODS: Fifty rabies virus isolates from bovines and equines in the States of São Paulo and Minas Gerais, Brazil, were genetically characterized and compared with sequences retrieved from GenBank. RESULTS: Two clusters (I and II) with mean nucleotide identities of 99.1 and 97.6 percent were found. The first of these contained nearly all the samples analyzed. Lineages from other Brazilian states grouped in cluster II. CONCLUSIONS: Analysis of the amino acid sequences of the N proteins revealed the existence of genetic markers that may indicate possible variations between geographic regions, although the biologically active regions are conserved within the species over space and time.

INTRODUÇÃO: A raiva é uma doença aguda do sistema nervoso central e é responsável por mortes de milhares de humanos, animais silvestres e animais de criação - especialmente bovinos - além de causar elevadas perdas econômicas. Este trabalho descreve a caracterização genética das variantes do vírus da raiva que circulam em populações de Desmodus rotundus e são transmitidas aos herbívoros. MÉTODOS: Cinquenta isolados de vírus da raiva de bovinos e equinos provenientes dos Estados de São Paulo e Minas Gerais, Brasil, foram caracterizadas geneticamente e comparadas com sequências recuperadas do GenBank. RESULTADOS: Dois clusters, I e II, apresentando identidades médias de nucleotídeos de 99,1 e 97,6 por cento, foram obtidos, sendo o primeiro composto de quase a totalidade das amostras analisadas. Linhagens de outros estados do Brasil "clustered" no II. CONCLUSÕES: A análise das sequências de aminoácidos da proteína N revelou que existem marcadores genéticos que podem determinar uma possível regionalidade embora as regiões biologicamente ativas apresentem-se conservadas dentro das espécies ao longo do tempo e espaço.

Genetic characterization of Rabies virus isolated from cattle between 1997 and 2002 in an epizootic area in the state of São Paulo, Brazil.

The biogeographical history of rabies can be reconstructed using molecular data. This work describes the genetic characterization of the Rabies virus variant that circulates in the Desmodus rotundus (vampire bat) population in an epizootic area and is transmitted to herbivorous livestock. The N and G genes of this virus were sequenced, and the phylogenetic trees generated were topologically concordant. Three genetic clusters were identified in the epizootic area and were designated RD1, RD2 and RD3. The results show that the origins of the epizootics in areas RD1 and RD2 were different and that the epizootic in area RD3 was the result of expansion of that in area RD2. The two genes analyzed are conserved, and their identities, which are greater than 98%, were maintained over time and space. The genetic sequences in this study were compared with others retrieved from GenBank, and the high identity of the N and G genes was also shown to be maintained over time and space. The results suggest that the D. rotundus lineages of the Rabies virus from the Atlantic coast of South America are highly conserved.

Molecular characterization of Rabies Virus isolates from dogs and crab-eating foxes in Northeastern Brazil.

Thirty-eight samples of Rabies Virus isolated from dogs and crab-eating foxes (Cerdocyon thous) in Northeastern Brazil were characterized genetically by analyzing the G gene and the psi region. The results show that there are two groups of Rabies Virus lineages circulating among domestic and wild animals in the region. The topologies of the phylogenetic trees of the G gene and psi region are similar and reveal the existence of geographic groups. The genetic diversity of the lineages isolated from wild animals (wild group) was approximately twice that of the lineages isolated from domestic animals (domestic group), and the genetic distance between the two groups was 9.93%. Polymorphism analysis revealed specific intra- and inter-group molecular signatures for both the G gene and psi region. Together with the analysis of the N gene undertaken previously, the results of this study confirm the existence of a Rabies Virus phylogroup in Northeastern Brazil (NB) circulating in the C. thous population, making this species a rabies biotype in the region.

Species determination of Brazilian mammals implicated in the epidemiology of rabies based on the control region of mitochondrial DNA

Identification of animals that are decomposing or have been run over or burnt and cannot be visually identified is a problem in the surveillance and control of infectious diseases. Many of these animals are wild and represent a valuable source of information for epidemiologic research as they may be carriers of an infectious agent. This article discusses the results obtained using a method for identifying mammals genetically by sequencing their mitochondrial DNA control region. Fourteen species were analyzed and identified. These included the main reservoirs and transmitters of rabies virus, namely, canids, chiroptera and primates. The results prove that this method of genetic identification is both efficient and simple and that it can be used in the surveillance of infectious diseases which includes mammals in their epidemiologic cycle, such as rabies.

A coronavirus detected in the vampire bat Desmodus rotundus

This article reports on the identification of a group 2 coronavirus (BatCoV DR/2007) in a Desmodus rotundus vampire bat in Brazil. Phylogenetic analysis of ORF1b revealed that BatCoV DR/2007 originates from a unique lineage in the archetypical group 2 coronaviruses, as described for bat species elsewhere with putative importance in Public Health.

Species determination of Brazilian mammals implicated in the epidemiology of rabies based on the control region of mitochondrial DNA.

Identification of animals that are decomposing or have been run over or burnt and cannot be visually identified is a problem in the surveillance and control of infectious diseases. Many of these animals are wild and represent a valuable source of information for epidemiologic research as they may be carriers of an infectious agent. This article discusses the results obtained using a method for identifying mammals genetically by sequencing their mitochondrial DNA control region. Fourteen species were analyzed and identified. These included the main reservoirs and transmitters of rabies virus, namely, canids, chiroptera and primates. The results prove that this method of genetic identification is both efficient and simple and that it can be used in the surveillance of infectious diseases which includes mammals in their epidemiologic cycle, such as rabies.

A coronavirus detected in the vampire bat Desmodus rotundus.

This article reports on the identification of a group 2 coronavirus (BatCoV DR/2007) in a Desmodus rotundus vampire bat in Brazil. Phylogenetic analysis of ORF1b revealed that BatCoV DR/2007 originates from a unique lineage in the archetypical group 2 coronaviruses, as described for bat species elsewhere with putative importance in Public Health.

Characterization of Rabies virus isolated from canids and identification of the main wild canid host in Northeastern Brazil.

The rabies cases in dogs and wild canids in Northeastern Brazil are a public and animal health problem. This paper describes the identities of the coding region of the N-gene of Rabies virus (RABV) isolated in canids from Northeastern Brazil. The genetic tree generated using the sequence data described here divided the cluster BRAZILAN CANIDS into two subclusters (DOG-RELATED STRAINS and WILD CANID-RELATED STRAINS) with identities greater than those already described. The two subclusters are sub-divided into geographic groups related to the origin of the isolates, suggesting a long-standing ecological coexistence of the sequence types characteristic of the groups. This article also analyzes the 513-nucleotide stretch of the mitochondrial DNA control region of rabies-positive canids from Northeastern Brazil with a view to identifying the main RABV host among them. Among the four species of wild canids found in the region, two (Cerdocyon thous and Pseudalopex vetulus) are frequently associated with rabies. Phylogenetic analysis of sequence data generated from mtDNA suggests that C. thous is the main wild canid host in the region. The results obtained in this study are in concordance with the zoology and ecology of wild canids, and thus, help improve epidemiologic vigilance of rabies and allow a more targeted control of the disease.