Lyssaviruses are bullet shaped negative-sense RNA viruses that are all able to cause the fatal encephalitic disease known as rabies. The genus currently consists of 17 formally recognised viral species with one tentative species awaiting classification. The prototype virus for the Lyssavirus genus is the well-known rabies virus (RABV), while all other species in the genus are classified as rabies-related viruses. In South Africa specifically, RABV, Lagos bat virus (LBV), Duvenhage virus (DUVV), and Mokola virus (MOKV) are known to circulate, with RABV and DUVV associated with human fatalities. Active surveillance on rabies-related lyssaviruses in bats, specifically African insectivorous bat species, is either very sporadic or non-existing, providing an inaccurate overall representation of prevalence, diversity, and geographic distribution. Therefore, we conducted viral nucleic acid surveillance for lyssaviruses in different insectivorous bat species in South Africa. These samples were collected during routine field surveillance and included bats that were found dead, appeared to be displaying abnormal behavior or taken as vouchers specimens as part of bat taxonomic studies. A quantitative real-time reverse transcription polymerase chain reaction assay, capable of detecting the diversity of lyssaviruses were used to test extracted RNA. Three brain samples tested positive and were further characterized by conventional RT-PCR, DNA sequencing and phylogenetic analyses targeting the nucleoprotein gene. One of the positive brains was detected from a Common slit-faced bat (Nycteris thebaica) and the other two positive brains were detected from the Natal long-fingered bat (Miniopterus natalensis). Phylogenetic analysis of the nucleoprotein indicated one detection to be a Duvenhage lyssavirus with the other two detections showing a close relationship with the West Caucasian bat virus species, previously only detected in Eastern Europe. However, a more than 20 % nucleotide divergence indicated it to be a potentially new lyssavirus species, Matlo bat lyssavirus. The virus was successfully isolated using the mouse inoculation test followed by full genome next generation amplicon sequencing. The results of the full genome characterisation further supported the initial findings with concatenated coding regions nucleotide divergence ranging between 16% and 23.7% as well as consistent phylogenetic tree topology groupings identical to initial phylogenetic analyses using multiple evolutionary models. The identification of a putative new lyssavirus highlights the importance of routine lyssavirus surveillance to understand the diversity. Further investigation is required to determine the possible reservoir species since the Natal long-fingered bats are known to co-roost with different bat species in caves. The potential of spillover to humans and other animals is unknown but people often enter these bat roosts for traditional and recreational purposes and bats do come into contact with several animal species including humans during foraging. / Dissertation (MSc)--University of Pretoria, 2021. / CDC- 5 NU2GGH001874-02-00 NRF-78566 / Medical Virology / MSc / Restricted

Dissertação (mestrado)—Universidade de Brasília, Faculdade de Agronomia e Medicina Veterinária, Programa de Pós-Graduação em Saúde Animal, 2014. / Submitted by Ana Cristina Barbosa da Silva (annabds@hotmail.com) on 2015-06-02T16:29:05Z No. of bitstreams: 1 2014_SileneManriqueRocha_Parcial.pdf: 581235 bytes, checksum: 06c91e2abfd9133d8817a8d59afad13c (MD5) / Approved for entry into archive by Raquel Viana(raquelviana@bce.unb.br) on 2015-06-22T21:03:34Z (GMT) No. of bitstreams: 1 2014_SileneManriqueRocha_Parcial.pdf: 581235 bytes, checksum: 06c91e2abfd9133d8817a8d59afad13c (MD5) / Made available in DSpace on 2015-06-22T21:03:34Z (GMT). No. of bitstreams: 1 2014_SileneManriqueRocha_Parcial.pdf: 581235 bytes, checksum: 06c91e2abfd9133d8817a8d59afad13c (MD5) / A raiva é uma das doenças transmissíveis mais importantes no mundo, devido ao elevado impacto em saúde pública e a maior taxa de mortalidade de todos agentes patógenos virais humanos. Estudos apontam a presença do Lyssavirus em reservatórios do ciclo silvestre evidenciando a participação de canídeos silvestres, saguis, morcegos hematófagos e não hematófagos entre os potenciais transmissores da doença para animais domésticos e humanos. Assim, a reintrodução da raiva no ambiente urbano a partir do ciclo silvestre é uma preocupação. Este estudo descreve o perfil de ocorrência de raiva no Brasil, de 2002 a 2012, com ênfase no ciclo de transmissão silvestre da doença. Foi realizado um estudo descritivo utilizando registros de tempo, pessoa e lugar dos casos confirmados em humanos e animais, como também de atendimentos antirrábicos registrados no banco de dados Sinan. No período, foram notificados 82 casos de raiva em humanos, transmitidos por animais silvestres, predominantemente nas regiões Norte e Nordeste em áreas rurais, com maior freqüência no sexo masculino. Observou-se que 72% dos casos humanos não receberam procedimentos de profilaxia pós-exposição (PEP). Dentre os mamíferos silvestres transmissores da doença, houve destaque para o morcego hematófago. Os 460 mamíferos silvestres terrestres confirmados com a doença foram notificados no Nordeste. No período houve registro de 1.703 morcegos infectados com vírus rábico e a região Sudeste se destacou pela frequência de registros de morcegos não hematófagos. Foi observado que as regiões Centro-Oeste e Norte apresentaram menor proporção de registros de raiva em mamíferos silvestres terrestres. O número elevado de casos de raiva em bovinos evidenciou, entretanto, a importância do morcego hematófago, como mantenedor do vírus rábico no ciclo rural. Os resultados apresentados são úteis para planejamento da vigilância da raiva no País. ____________________________________________________________________________________ ABSTRACT / Rabies is one of the most important communicable diseases in the world, due to high impact on public health and mortality approaches 100%. Studies indicate the presence of Lyssavirus in reservoirs showing the sylvatic cycle involving wild canids, marmosets, vampire bats and nonhematophagous as potential transmitters of disease to animals and humans. Thus, the reintroduction of rabies in the urban environment from the wildlife is a concern. This study describes the profile of the occurrence of rabies in Brazil, in the period 2002-2012, with emphasis on the wildlife transmission cycle of the disease. A descriptive study using the database Sinan data records with time approach, person and place of confirmed cases in humans and animals, as well antirrábico care was conducted. 84 cases of rabies in humans have been reported, predominantly in the North and Northeast regions, in rural areas, more frequently in males and 80 % of these did not receive prophylaxis. Among the wild species, the vampire bat is the largest transmitter of the disease to humans, the same was observed in antirrábicos care in the North. Is 460 terrestrial wild mammals confirmed with the disease were reported in northeast. 1703 bats, especially the vampire bats with no greater proportion of notifications in the Southeast were recorded. The North and Midwest regions showed no reported cases of rabies in wild mammals. The number of cattle was significant, showing that there was presence of vampire bat rabies virus that keeps rural disease cycle. Thus the results may be useful for surveillance of rabies in the country.

Les virus zoonotiques sont une menace pour les humains à cause de leur capacité à passer des réservoirs animaux à l'homme. La rage est provoquée par un virus zoonotique (Virus de la Rage) qui a de multiples réservoirs animaux. La rage est contractée lors de la morsure par un animal infecté et est incurable et létale après l'apparition des premiers symptômes. Un défi visant à éradiquer la rage chez les chiens à l'horizon 2030 a été proposé. Il imposera de stopper la transmission du virus aux populations de chiens à partir des autres réservoirs animaux. Pour cela, il est essentiel de comprendre les mécanismes de transmission entre hôtes potentiels du virus. Dans notre thèse, nous faisons l'hypothèse que le passage d'un hôte à un autre est lié à la diversité des populations virales chez un hôte donné, appelée "hétérogénéité virale".Pour étudier cette hétérogénéité virale, des méthodes de séquençage des populations virales ont été développées. La transmission du virus de la rage entre chiens a été analysée et un évènement de transmission entre chien et renard a été étudié. Une plus importante hétérogénéité virale a été observée chez le renard après sa contamination par le chien en comparaison avec d'autres renards infectés par des congénères de la même espèce. Ceci suggère que l'hétérogénéité virale est importante dans le phénomène de transmission inter-espèce. Ces résultats sont importants pour améliorer notre compréhension de l'évolution du virus de la rage chez un nouvel hôte et pourront aider les efforts d'éradication de la maladie. / Zoonotic viruses are a threat to humans, jumping from animal reservoirs into humans. Rabies is caused by rabies virus (RABV), a zoonotic virus, with many animal reservoirs. Rabies is contracted from a bite of infected animal and once symptoms appear, death is inevitable. A challenging target date of 2030 to eliminate rabies in dogs has been set. One challenge will be stopping RABV re-entering the dog population from other animal reservoirs. Understanding how RABV switches hosts is important to prevent it happening. In this thesis, I hypothesise that successful host switching is due to the diverse population of viruses within the host termed ‘viral heterogeneity’. To investigate viral heterogeneity, methods to sequence the virus populations within clinical samples were developed. Transmission of RABV within dogs was analysed and a host shift event from dogs to foxes was investigated. High viral heterogeneity was seen in foxes after the host shift than in other foxes, suggesting it is important for a successful host shift. These data will be important to improve our understanding of how viruses evolve in new hosts, helping governments to eradicate disease.

Mokola virus (MOKV), a rabies-related lyssavirus, represents one of 17 recognized species within the Lyssavirus genus, all of which are capable of causing the fatal encephalitic rabies disease. MOKV is exclusively endemic to Africa with only 30 sporadic cases reported since its discovery more than 50 years ago. The reservoir host for MOKV remains unknown, however, several hypotheses have been formulated. Small non-volant mammals (i.e. shrews, sengis and rodents) have been suggested as possible reservoir hosts with previous MOKV isolations from shrews (Crocidura spp.) and a single rodent (Lophuromys sikapusi) providing support of the first lyssavirus species that has an association with small non-volant mammals. To investigate further, nucleic acid- and serological surveillance were conducted in small non-volant mammals from Southern Africa (specifically South Africa and Mozambique). Nucleic acid surveillance with a pan-lyssavirus qRT-PCR assay of 355 brain samples did not identify any new MOKV infections. Serological surveillance using a micro-neutralization test of 287 serum samples identified 37 samples that were positive for the presence of MOKV virus neutralizing antibodies. These positive serum samples indicate previous MOKV exposure and were all collected from Bushveld gerbils (Gerbilliscus leucogaster) in both South Africa (n=36) and Mozambique (n=1). From all of the Bushveld gerbils that were tested, an overall MOKV seropositivity of 87.80% is observed for the gerbils that were caught at Meletse in Limpopo. Since MOKV have been shown to cross-react in serological assay with closely-related lyssaviruses, the seropositivity observed could have been due to exposure of another phylogroup II lyssavirus. Serological evidence of MOKV in this rodent species was previously observed in a study conducted in Zimbabwe in 1988, which raises their profile as a potential MOKV host candidate. Experimental pathogenicity studies support this notion due to significant amounts of MOKV found in their salivary glands that could be sufficient for transmission. To gain further insight of the phylogeny and genetic diversity of MOKV, complete genome sequences of three historic MOKV isolates from South Africa (MOKV 700/70, 229/97, and 12/458) were generated. Future studies are needed to expand surveillance, detection and characterization of lyssaviruses. / Dissertation (MSc (Medical Virology))--University of Pretoria, 2020. / The Centre for Disease Control and Prevention (CDC) Global Disease Detection (GDD) Programme (Corporate Agreement Number: 5 NU2GGH001874-02-00). / The South African Research Chair in Infectious Diseases of Animals (Zoonoses) from the National Research Foundation (NRF) of the Department of Science and Innovation (DSI), Prof. Wanda Markotter (UID98339), as well as additional grants awarded to Prof. Markotter by the NRF (UID92524, UID85756 & UID91496). / Medical Virology / MSc (Medical Virology) / Restricted

In Chapter 1, the literature relating to rabies virus and the rabies like lyssaviruses is reviewed. In Chapter 2 data are presented from 1170 diagnostic submissions for ABLV testing by fluorescent antibody test (Centocor FAT). All 27 non-bat submissions were ABLV-negative. Of 1143 bat accessions 74 (16%) were ABLV-positive, including 69 of 974 (7.1%) flying foxes (Pteropus spp.), 5 of 7 (71.4%) Saccolaimus flaviventris (Yellow-bellied sheathtail bats), none of 151 other microchiropteran bats, and none of 11 unidentified bats. Statistical analysis of data from 868 wild Black, Grey-headed, Little Red and Spectacled flying foxes (Pteropus alecto, P. poliocephalus, P. scapulatus, and P. conspicillatus) indicated that three factors; species, health status and age were associated with significant (p&lt 0.001) differences in the proportion of ABLV-positive bats. Other factors including sex, whether the bat bit a person or animal, region, year, and season submitted, were not associated with ABLV. Case data for 74 ABLV-positive bats, including the circumstances in which they were found and clinical signs, is presented. In Chapter 3, the aetiological diagnosis was investigated for 100 consecutive flying fox submissions with neurological signs. ABLV (32%), spinal and head injuries (29%), and neuro-angiostrongylosis (18%) accounted for most neurological syndromes in flying foxes. No evidence of lead poisoning was found in unwell (n=16) or healthy flying foxes (n=50). No diagnosis was reached for 16 cases, all of which were negative for ABLV by TaqMan® PCR. The molecular diversity of ABLV was examined in Chapter 4 by sequencing 36 bases of the leader sequence, the entire N gene, and start of the P gene of 28 isolates from pteropid bats and 3 isolates from Yellow-bellied sheathtail (YBST) bats. Phylogenetic analysis indicated all ABLV isolates clustered together as a discrete group within the Lyssavirus genera closely related to rabies virus and European bat lyssavirus-2 isolates. The ABLV lineage consisted of two variants; one (ybst-ABLV) consisted of isolates only from YBST bats, the other (pteropid-ABLV) was common to Black, Grey-headed and Little Red flying foxes. No associations were found between the sequences and either the geographical location or year found, or individual flying fox species. In Chapter 5, 15 inocula prepared from the brains or salivary glands of naturally-infected bats were evaluated by intracerebral (IC) and footpad (FP) inoculation of Quackenbush mice in order to select and characterize a highly virulent inoculum for further use in bats (Inoculum 5). In Chapter 6, nine Grey-headed flying foxes were inoculated with 105.2 to 105.5 MICED50 of Inoculum 5 divided into four sites, left footpad, pectoral muscle, temporal muscle and muzzle. Another bat was inoculated with half this dose divided into the footpad and pectoral muscle only. Seven of 10 bats developed clinical disease of 1 to 4 days duration between PI-days 10 and 19 and were shown to be ABL-positive by FAT, HAM immunoperoxidase staining, virus isolation in v mice, and TaqMan PCR. Five of the seven bats displayed overt aggression, one died during a seizure, and one showed intractable agitation, pacing, tremors, and ataxia. Viral antigen was demonstrated throughout the central and peripheral nervous systems and in the epithelial cells of the submandibular salivary glands (n=4). All affected bats had mild to moderate non-suppurative meningoencephalitis and severe ganglioneuritis. No ABLV was detected in three bats that remained well until the end of the experiment on day 82. One survivor developed a strong but transient antibody response. In Chapter 7, the relative virulence of inocula prepared from the brains and salivary glands of experimentally infected flying foxes was evaluated in mice by IC and FP inoculation and TaqMan assay. The effects in mice were correlated to the TaqMan CT value and indicated a crude association between virulence and CT value that has potential application in the selection of inocula. In Chapter 8, 36 Black and Grey-headed flying foxes were vaccinated with one (day 0) or two (+ day 28) doses of Nobivac rabies vaccine and co-vaccinated with keyhole limpet haemocyanin (KLH). All bats responded to the Nobivac vaccine with a rabies-RFFIT titer &gt 0.5 IU/mL that is nominally indicative of protective immunity. Plasma from bats with rabies titres &gt 2 IU/mL had cross-neutralising ABLV titres &gt 1:154. A specifically developed ELISA detected a strong but transient response to KLH.

Duvenhage virus (DUVV) belongs to genotype (gt) 4 of the lyssavirus genus, in the family Rhabdoviridae, order Mononegavirales. This virus causes fatal rabies encephalitis and has only been reported from the African continent. To date there have been only five isolations of DUVV, three of which were from human fatalities and all of which were associated with insectivorous bat species. Genotype 4 lyssaviruses have not been well studied and as such little is known about them. The aim of this study was to determine the full genome sequence and investigate the epidemiology of this uniquely African lyssavirus. Standard methods of PCR and sequencing were used to determine the coding and non coding regions of various DUVV isolates. In order to determine the full genome sequence, an RNA circularization technique was used to obtain the genomic terminal sequences. Using various molecular techniques we then analyzed the sequence data, at both phylogenetic and evolutionary levels. Our analysis showed the evolutionary forces acting against DUVV, to be similar to that which has been found for its closest relative, European bat lyssavirus type 1 (EBLV1) (gt 5). Both these viruses have strong constraints against amino acid change, with no evidence of positive selection. Phylogenetic studies showed that not all Lyssavirus genes are equal for phylogenetic or lyssavirus classification analysis. High intergenotypic values at the nucleoprotein amino acid level emphasize that there is a need to reinvestigate the criteria for lyssavirus genotype classification. The strong support observed in our full genome studies suggests that full genomes may in fact be best for Lyssavirus analysis, so as to avoid the potential bias of individual gene analyses. Analysis of DUVV indicates that it is an older virus within the lyssavirus genus and as shown by the discovery of the most recent isolate, the genetic diversity and incidence of this virus is greatly underestimated. Poor surveillance of rabies-related lyssaviruses as well as the poor diagnostic capabilities through most of Africa are large contributors to our lack of information. Improved surveillance of the African rabies-related lyssaviruses will extend our knowledge on the geographic distribution, host species associations and epidemiology of these viruses. / Dissertation (MSc)--University of Pretoria, 2011. / Microbiology and Plant Pathology / unrestricted

Submitted by GISLAINE RAQUEL SANTOS (g_raquelsantos@yahoo.com.br) on 2016-07-12T13:44:01Z No. of bitstreams: 1 Tese CARACTERIZAÇÃO EPIDEMIOLÓGICA E MOLECULAR DA RAIVA EM BOVINOS NO ESTADO Gislaine Raquel dos Santos.pdf: 3457607 bytes, checksum: 5f07e27a43061393bdefd203b8570a11 (MD5) / Approved for entry into archive by Ana Paula Grisoto (grisotoana@reitoria.unesp.br) on 2016-07-14T18:07:47Z (GMT) No. of bitstreams: 1 santos_gr_dr_jabo.pdf: 3457607 bytes, checksum: 5f07e27a43061393bdefd203b8570a11 (MD5) / Made available in DSpace on 2016-07-14T18:07:47Z (GMT). No. of bitstreams: 1 santos_gr_dr_jabo.pdf: 3457607 bytes, checksum: 5f07e27a43061393bdefd203b8570a11 (MD5) Previous issue date: 2016-03-22 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / A raiva é uma antropozoonose de evolução letal causada por vírus do gênero Lyssavirus. É uma das doenças infecciosas responsáveis por causar prejuízos aos produtores rurais, levando a impactos econômicos significativos no agronegócio. Objetivou-se com o presente trabalho determinar o perfil epidemiológico da raiva em herbívoros no Estado de Pernambuco, Brasil, no período de 2007 a 2012. Foi realizado um estudo retrospectivo dos dados relativos aos casos positivos de raiva de herbívoros, levando em consideração o mês e o ano da ocorrência e a região geográfica. As análises moleculares foram desenvolvidas a partir de amostras de encéfalos provenientes das cinco Mesorregiões (Agreste, Mata, Sertão, Metropolitana e São Francisco) do Estado. No período estudado foram detectados 238 resultados positivos para o vírus da raiva em herbívoros, distribuídas espacialmente nas cinco mesorregiões, em 78 (42,1%) dos 185 municípios. Observou-se no decorrer do período uma diminuição significativa na taxa de incidência, com ausência de sazonalidade. Quando se analisou a taxa de incidência levando em consideração as Mesorregiões, observou-se que a Mata foi a que apresentou maior oscilação. Para complementar a análise epidemiológica, 16 amostras foram submetidas à técnica de RT-PCR para amplificação parcial do gene N. As sequências geradas foram alinhadas com sequências homólogas obtidas no GenBank para a construção da árvore filogenética, pelo método Bayesiana. Todas as amostras foram homólogas às sequências de vírus da raiva relacionadas à linhagem do morcego hematófago Desmodus rotundus. Diante dos resultados obtidos, constata-se que o vírus da raiva está presente em todo o Estado de Pernambuco, relacionado à linhagem do morcego hematófago Desmodus rotundus. Os focos de raiva em herbívoros estão distribuídos em graus diferenciados em todas as mesorregiões, porém não se constatou um aumento no número de casos que se repita de forma sistemática em uma mesma época do ano, indicando ausência de sazonalidade. Observou-se, também, uma significativa diminuição da incidência no decorrer do período estudado. Esse panorama enfatiza a importância da contínua realização das atividades de prevenção e controle pela Vigilância Agropecuária. / Rabies is an anthropozoonosis with lethal evolution caused by genus lyssavirus viruses. It is one among infectious diseases who are responsible for causing losses to farmers, leading to a significant economic impacts on agribusiness. The objective of this study was to determine the epidemiological profile in herbivores whithin the State of Pernambuco, Brazil, from 2007 to 2012. A retrospective study was conducted based on data from positive cases of rabies of herbivores, considering the month, the year of occurrence and geographic region. The molecular analyzes were developed using brain samples from the five Mesoregions (Agreste, Mata, Sertão, Metropolitana and São Francisco) within the State. During the study period, were detected positive for rabies 238 from herbivores, spatially distributed in 78 (42.1%) of 185 municipalites from the five mesoregions. It was observed, during the period cited, a significant decrease in the incidence rate, with no seasonal nature. While analyzing the incidence rate considering the mesoregions, the Mata region showed the greatest oscillation. As a complement to the epidemiological analysis, 16 samples were subjected to RT-PCR for the partial amplification of the gene N. The generated sequences were aligned with homologous sequences obtained from the GenBank to build the phylogenetic tree by bayesian method. On the results, it was found that the rabies virus is present all over the state of Pernambuco, related to the lineage of vampire bat Desmodus rotundus. The rabies outbreaks in herbivores are distributed in different degrees throughout the mesoregions but it was not found an increase in cases numbers that repeats systematically during the same time of the years, indicating, therefore, the absence of seasonality. It was observed also a significant decrease in incidence over the studied period. This scenario emphasizes the importance of continued prevention and control activities by the Agricultural Surveillance.

Submitted by Rebeka Godeiro (rebeka_carvalho@hotmail.com) on 2018-05-21T13:16:05Z No. of bitstreams: 1 ANA CLÁUDIA CAETANO PINHEIRO LEITE - DISSERTAÇÃO ZOOTECNIA 2017.pdf: 1967513 bytes, checksum: f63f25f8174ca192203382662c1a5ffc (MD5) / Made available in DSpace on 2018-05-21T13:16:05Z (GMT). No. of bitstreams: 1 ANA CLÁUDIA CAETANO PINHEIRO LEITE - DISSERTAÇÃO ZOOTECNIA 2017.pdf: 1967513 bytes, checksum: f63f25f8174ca192203382662c1a5ffc (MD5) Previous issue date: 2017-05-31 / Capes / Esta dissertação é composta por dois capítulos. No Capítulo I, foi realizada uma revisão de literatura, que enfocou a raiva como uma zoonose que permeia vários ciclos epidemiológicos. Os morcegos são animais que possuem uma variedade de hábitos alimentares e integrados a eles importantes funções ecológicas. Entretanto os mais conhecidos estão associados a interações negativas, pois se alimentam de sangue (hematófagos) de aves e mamíferos. A espécie Desmodus rotundus, se alimenta estritamente de sangue de mamíferos e hoje é a principal responsável pela veiculação do vírus na região rural, principalmente para bovinos. As modificações nos ambientes naturais, inclusive no bioma Caatinga, que ocupa quase todo território paraibano, desencadeiam desequilíbrios na natureza ativando zoonoses ―adormecidas‖ como a raiva. Para impedir o ressurgimento de doenças que envolvem vários ciclos devem se trabalhar mecanismos de previsibilidade de zonas de risco. Ferramentas de Geoprocessamento, como SIG (Sistema de Informação Geográfica) pode estimar áreas vulneráveis ao vírus, através da espacialização de dados georreferenciados sobre focos e locais de convívio com o principal transmissor. No Capítulo II, foi desenvolvido um trabalho com o objetivo de caracterizar espacialmente abrigos de morcegos e ocorrências da raiva na região semiárida da Paraíba. Foram utilizados dados fornecidos pela Secretaria de Estado do Desenvolvimento da Agropecuária e da Pesca da Paraíba (SEDAP) do período de 2007 a 2015, informações coletadas da plataforma digital do IBGE (Instituto Brasileiro de Geografia e Estatística) e um mosaico de imagens do INPE (Instituto Nacional de Pesquisas Espaciais). As representações geográficas foram produzidas através do software QGIS 2.16.0 – Nodebo, com malhas cartográficas do IBGE e da Agência Executiva de Gestão das Águas do Estado da Paraíba. No período de 2007 a 2015 foram cadastrados 93 abrigos e registradas 51 ocorrências de raiva distribuídas em todas as mesorregiões do Estado. Através dos abrigos georreferenciados foram plotados buffers de 10 km para verificar os locais de maior risco de circulação do vírus da raiva. Considerando o raio de 10 km mais de 80 municípios estão dentro da zona de risco da circulação do vírus rábico, além de outras cidades de Pernambuco, Ceará e Rio Grande do Norte, que ficam na divisa da Paraíba. As notificações isoladas, distantes de abrigos cadastrados, demonstram a necessidade de expandir os trabalhos de identificação de abrigos de morcegos, principalmente com a presença de D. rotundus. As subnotificações das ocorrências da raiva no Estado evidenciam que as autoridades 9 competentes devem elucidar melhor os proprietários de informar sobre o surgimento de doenças nervosas em seus rebanhos, como também aumentar o número de informações sobre a presença do D. rotundus na região. / This Master’s thesis comprises two chapters. Chapter 1 presents a literature review that focuses on rabies as a zoonosis that undergoes several epidemiological cycles. Bats are animals with a variety of feeding habits which are directly related to important ecological functions. However, the best known ones are associated to negative interactions for being hematophagous (feed on the blood of birds and mammals). The species Desmodus rotundus, feeds strictly on the blood of mammals and it is currently considered the number one responsible for spreading the rabies virus in rural regions, especially on cattle. The changes in the natural environments, including the Caatinga biome, which stretches throughout most of the Paraíba state territory in Brazil, caused unbalances in the nature, thus reactivating zoonosis that had been inoperative, such as rabies. In order to impede the resurgence of diseases that involve various cycles, mechanisms of predictability in risks zones must be employed. Geoprocessing tools such as the SIG (Brazilian acronym for Geographic Information System) might estimate the vulnerable areas to the virus through spacing of georreferenced data of the sites and places of conviviality with the illness-transmitting animals. Chapter 2 describes an experience conducted with the objective of characterizing spatially the bats’ shelters and the events of rabies in the semi-arid regions (droughty lands) of Paraíba. This work included data from the SEDAP (Brazilian acronym for Paraíba Farming and Ranching Activities Development State Department) between 2007 and 2015; as well as information from the IBGE (Brazilian acronym for Brazilian Institute of Geography and Statistics) digital platform; and even a mosaic image of the INPE (Brazilian acronym for National Institute for Space Research). The geographical representations were produced on QGIS 2.16.0 – Nodebo software with a cartographic mesh from the IBGE and Paraíba State Executive Agency of Water Management. Over the period between the years of 2007 and 2015, 93 bat shelters and 51 occurrences of rabies were catalogued at all state’s mesoregions. Based on the georeferenced shelters, 10-kilometer-buffers were plotted to verify the sites of larger risk of circulation of the rabies virus. Within a 10-kilometer-radius, over 80 towns are in the risk zone of the virus circulation as well as a few other municipalities from the neighboring states of Pernambuco, Ceará and Rio Grande do Norte that are located right on their states’ borders. Isolated cases of rabies infections far from the catalogued shelters show the need for expanding the efforts of identifying the bats’ shelters especially the ones where D. rotundus are present. The undernotes of rabies infections within the state 11 demonstrate that government authorities must provide better information to the farmers about the raise of nerve diseases in the cattle, as well as increase the amount of notifications of the presence of D. rotundus in the region.

La rage est une zoonose mortelle, susceptible d’atteindre autant les mammifères sauvages et domestiques que l’Homme. Elle est à l’origine d’environ 70 000 décès humain déclarés par an, majoritairement des enfants dans les pays en développement. Le chien, réservoir majeur de l’espèce RABV, est à l’origine de 98-99% de ces décès. Quatorze espèces de Lyssavirus, circulant majoritairement chez les chiroptères sont actuellement reconnues. La vaccination, associée à des mesures sanitaires, reste le meilleur outil de prévention et de maîtrise de la maladie. A l’heure actuelle, seule la sérologie permet de contrôler l’efficacité de la vaccination antirabique, le développement des anticorps neutralisants étant le premier témoin d’une immunité protectrice. Les travaux s’appuyant sur la séroneutralisation virale, et notamment ceux auxquels j’ai participé, ont mis en évidence l’influence de différents facteurs dont certains ont conduit à préconiser des modifications de protocoles vaccinaux. Ils ont également permis d’assurer le suivi de l’efficacité de la vaccination individuelle ou de groupe sur le terrain et de contribuer à son amélioration. Les tests de séroneutralisation sont également utilisés dans le cadre de l’épidémiosurveillance de populations animales non vaccinées. La mise en œuvre de ces tests chez les chiroptères en France, après leur adaptation au Lyssavirus d’intérêt que j’ai menée à bien, a permis d’obtenir des informations sur la circulation des espèces virales EBLV-1 et EBLV-2, sur une base uniquement sérologique pour ce dernier. D’autre part, elle a permis de mettre en évidence au sein d’une même colonie des phénomènes de transition sérologique au cours du temps, dont l’étude mériterait d’être approfondie. Les tests de séroneutralisation sont cependant difficilement transférables aux pays où la rage est très présente, du fait de ressources limitées. Mes travaux, proposant l’utilisation d’un test ELISA comme méthode alternative, ont contribué à remettre en cause le dogme du recours nécessaire à la séroneutralisation. Ce test, couplé à un système de collecte d’échantillons sanguins adapté au terrain, devrait améliorer le suivi de l’efficacité des campagnes de vaccination de la faune sauvage comme des animaux domestiques, y compris dans les pays d’enzootie où la qualité des prélèvements de sang ne peut être assurée. Ainsi, les outils d’évaluation de la réponse immunitaire humorale sont des outils très précieux au service de la lutte et de la surveillance de la rage animale dans le monde. Mes travaux, complémentaires à ceux réalisés par d’autres équipes, ont contribué à rendre envisageable l’objectif prioritaire des organisations internationales : l’éradication de la rage canine dans le monde à l’horizon 2030. Il est cependant nécessaire de les poursuivre pour améliorer les outils disponibles et d’en proposer de plus adaptés, afin d’atteindre l’ensemble des objectifs d’éradication, de la rage canine comme de la rage selvatique / Rabies is a deadly zoonosis that can affect wild and domestic mammals as much as humans. About 70,000 human deaths are reported each year, mostly in children from developing countries. Dogs, which are the major reservoir and source of the RABV species, account for 98-99% of these deaths. Currently, fourteen species of Lyssavirus, mainly circulating in chiroptera, are officially recognized. Vaccination, combined with sanitary measures, remains the best tool for preventing and controlling the disease. To date, only serology has allowed to control the effectiveness of rabies vaccination, as the production of neutralizing antibodies is the first evidence of protective immunity. Studies based on viral seroneutralisation, including my own studies, have highlighted the influence of various factors. Some of them have led to recommend modifications of vaccine protocols. They also contributed to monitor the effectiveness of individual or group vaccination field programmes and to improve these programmes. Seroneutralisation tests are also used in the context of the epidemiological surveillance of unvaccinated animal populations. I first successfully adapted these tests to lyssaviruses of interest in France. In a second step, their implementation in chiropters in France provided information on the circulation of EBLV-1 and EBLV-2 species, (only on a serological basis for the latter). This survey also allowed to highlight, within a specific colony, a phenomenon of serological transition over time, which should deserve to be studied further. However, seroneutralisation tests are difficult to be implemented in countries where rabies is very prevalent, mainly because of limited resources. My work, which recommends the use of an ELISA test as an alternative method, contributed to questioning the dogma of the necessary use of seroneutralisation tests. This test, coupled with a blood sampling system adapted to the field, should improve the monitoring of the effectiveness of vaccination campaigns for both wildlife and domestic animals, including in enzootic countries, where the quality of the blood samples cannot be guaranteed. Humoral immune response assessment tools are very valuable tools for the control and surveillance of animal rabies all around the world. My work, complementary to those carried out by other teams, has helped to make the priority objective of international organizations possible, i.e. the eradication of canine rabies in the world by 2030. However, further works are needed to improve the available tools and to propose more adapted ones, in order to achieve all the goals of eradication, for both canine and sylvatic rabies

Master of Public Health / Department of Diagnostic Medicine and Pathobiology / M. M. Chengappa / Rabies is the most deadly disease on earth and has a 99.9% human fatality rate. Rabies kills 61,000 humans annually and results in an economic burden of $124 billion USD annually. Each day 3.3 million people live with the risk of rabies. It is estimated that 95% of human rabies cases are a result of coming in contact with an infected canine, majority of these cases being children 15 years and younger. It is estimated that 1 person every 8 minutes dies of rabies. Rabies is a highly neurotropic disease which attacks the brain and central nervous system. Once clinical symptoms are presented, death is invariably the outcome as no cure exists for rabies. Rabies is 100% preventable in humans by proper wound management and proper administration of prophylaxis. Rabies can be adequately controlled in animal populations by contraception and animal rabies vaccine efforts. Whilst it is known that rabies can be prevented in humans and controlled in animal populations, further scientific efforts are still warranted to fully understand this deadly virus so that a cure can one day be discovered. As human and animal populations continue to grow, so does the cost and burden of this horrific disease. As a result, the importance of prophylaxis and passive immunity are critical in the event of medically managing an exposure, and preventing exposures. The World Health Organization has defined global recommendations for individuals and animals who have received prophylaxis to be adequately protected. Measuring this protection is performed using a variety of approved testing methodologies, virus-neutralizing assays and antigen-binding assays. Whilst the WHO recommendations were defined from clinical studies performed with virus-neutralizing assays, the assumption that these recommendations are suitable for the antigen-binding assays is inaccurate. The testing methodologies, virus-neutralization and antigen-binding, share similarities, as they are measuring an immune response to the rabies virus. However; enough differing characteristics are presented such that exact comparisons cannot be made. Establishing the same standards and recommendations for both testing methodologies will never be sufficient.