Rabies Anterograde Monosynaptic Tracing Allows Identification of Postsynaptic Circuits Receiving Distinct Somatosensory Input.

Somatosensory neurons detect vital information about the environment and internal status of the body, such as temperature, touch, itch, and proprioception. The circuit mechanisms controlling the coding of somatosensory information and the generation of appropriate behavioral responses are not clear yet. In order to address this issue, it is important to define the precise connectivity patterns between primary sensory afferents dedicated to the detection of different stimuli and recipient neurons in the central nervous system. In this study we describe and validate a rabies tracing approach for mapping mouse spinal circuits receiving sensory input from distinct, genetically defined, modalities. We analyzed the anatomical organization of spinal circuits involved in coding of thermal and mechanical stimuli and showed that somatosensory information from distinct modalities is relayed to partially overlapping ensembles of interneurons displaying stereotyped laminar organization, thus highlighting the importance of positional features and population coding for the processing and integration of somatosensory information.

A spatial and cellular distribution of rabies virus infection in the mouse brain revealed by fMOST and single-cell RNA sequencing.

BACKGROUND: Neurotropic virus infection can cause serious damage to the central nervous system (CNS) in both humans and animals. The complexity of the CNS poses unique challenges to investigate the infection of these viruses in the brain using traditional techniques. METHODS: In this study, we explore the use of fluorescence micro-optical sectioning tomography (fMOST) and single-cell RNA sequencing (scRNA-seq) to map the spatial and cellular distribution of a representative neurotropic virus, rabies virus (RABV), in the whole brain. Mice were inoculated with a lethal dose of a recombinant RABV encoding enhanced green fluorescent protein (EGFP) under different infection routes, and a three-dimensional (3D) view of RABV distribution in the whole mouse brain was obtained using fMOST. Meanwhile, we pinpointed the cellular distribution of RABV by utilizing scRNA-seq. RESULTS: Our fMOST data provided the 3D view of a neurotropic virus in the whole mouse brain, which indicated that the spatial distribution of RABV in the brain was influenced by the infection route. Interestingly, we provided evidence that RABV could infect multiple nuclei related to fear independent of different infection routes. More surprisingly, our scRNA-seq data revealed that besides neurons RABV could infect macrophages and the infiltrating macrophages played at least three different antiviral roles during RABV infection. CONCLUSION: This study draws a comprehensively spatial and cellular map of typical neurotropic virus infection in the mouse brain, providing a novel and insightful strategy to investigate the pathogenesis of RABV and other neurotropic viruses.

Inflammatory response and MAPK and NF-κB pathway activation induced by natural street rabies virus infection in the brain tissues of dogs and humans.

BACKGROUND: Street rabies virus (RABV) usually infects hosts at peripheral sites and migrates from motor or sensory nerves to the central nervous system. Several studies have found that inflammation is mild in a mouse model of street RABV infection. However, the pathogenetic mechanisms of street RABV in naturally infected dogs or humans are not well understood. METHODS: Brain tissues collected from 3 dogs and 3 humans were used; these tissue samples were collected under the natural condition of rabies-induced death. The inflammatory response and pathway activation in the brain tissue samples of dogs and humans were evaluated by HE, IHC, ARY006, WB and ELISA. The clinical isolate street RABV strains CGS-17 and CXZ-15 from 30 six-week-old ICR mice were used to construct the mouse infection model presented here. RESULTS: Neuronal degeneration and increased lymphocyte infiltration in the cerebral cortex, especially marked activation of microglia, formation of glial nodules, and neuronophagy, were observed in the dogs and humans infected with the street RABV strains. The various levels of proinflammatory chemokines, particularly CXCL1, CXCL12, CCL2, and CCL5, were increased significantly in the context of infection with street RABV strains in dogs and humans in relation to healthy controls, and the levels of MAPK and NF-κB phosphorylation were also increased in dogs and humans with natural infection. We also found that the degrees of pathological change, inflammatory response, MAPK and NF-κB signaling pathway activation were obviously increased during natural infection in dogs and humans compared with artificial model infection in mice. CONCLUSION: The data obtained here provide direct evidence for the RABV-induced activation of the inflammatory response in a dog infection model, which is a relatively accurate reflection of the pathogenic mechanism of human street RABV infection. These observations provide insight into the precise roles of underlying mechanisms in fatal natural RABV infection.

Adolescent frontal top-down neurons receive heightened local drive to establish adult attentional behavior in mice.

Frontal top-down cortical neurons projecting to sensory cortical regions are well-positioned to integrate long-range inputs with local circuitry in frontal cortex to implement top-down attentional control of sensory regions. How adolescence contributes to the maturation of top-down neurons and associated local/long-range input balance, and the establishment of attentional control is poorly understood. Here we combine projection-specific electrophysiological and rabies-mediated input mapping in mice to uncover adolescence as a developmental stage when frontal top-down neurons projecting from the anterior cingulate to visual cortex are highly functionally integrated into local excitatory circuitry and have heightened activity compared to adulthood. Chemogenetic suppression of top-down neuron activity selectively during adolescence, but not later periods, produces long-lasting visual attentional behavior deficits, and results in excessive loss of local excitatory inputs in adulthood. Our study reveals an adolescent sensitive period when top-down neurons integrate local circuits with long-range connectivity to produce attentional behavior.

Novel role of SARM1 mediated axonal degeneration in the pathogenesis of rabies.

Neurotropic viral infections continue to pose a serious threat to human and animal wellbeing. Host responses combatting the invading virus in these infections often cause irreversible damage to the nervous system, resulting in poor prognosis. Rabies is the most lethal neurotropic virus, which specifically infects neurons and spreads through the host nervous system by retrograde axonal transport. The key pathogenic mechanisms associated with rabies infection and axonal transmission in neurons remains unclear. Here we studied the pathogenesis of different field isolates of lyssavirus including rabies using ex-vivo model systems generated with mouse primary neurons derived from the peripheral and central nervous systems. In this study, we show that neurons activate selective and compartmentalized degeneration of their axons and dendrites in response to infection with different field strains of lyssavirus. We further show that this axonal degeneration is mediated by the loss of NAD and calpain-mediated digestion of key structural proteins such as MAP2 and neurofilament. We then analysed the role of SARM1 gene in rabies infection, which has been shown to mediate axonal self-destruction during injury. We show that SARM1 is required for the accelerated execution of rabies induced axonal degeneration and the deletion of SARM1 gene significantly delayed axonal degeneration in rabies infected neurons. Using a microfluidic-based ex-vivo neuronal model, we show that SARM1-mediated axonal degeneration impedes the spread of rabies virus among interconnected neurons. However, this neuronal defense mechanism also results in the pathological loss of axons and dendrites. This study therefore identifies a potential host-directed mechanism behind neurological dysfunction in rabies infection. This study also implicates a novel role of SARM1 mediated axonal degeneration in neurotropic viral infection.

Proteomics analysis of suckling mouse brain infected with attenuated rabies virus strain SRV9.

Rabies virus is an enveloped negative-stranded RNA virus belonging to the family Rhabdoviridae. It can be successfully controlled by vaccination however, there are still tens of thousands of deaths each year caused by rabies virus due to its mutations and complexity. A better understanding of the interaction between the rabies virus and the host might help solve this problem. Therefore, in this study, we used two-dimensional electrophoresis to investigate the protein expression of rabies virus-infected mice. This can help us to understand the impact of rabies virus on host protein expression during infection. For our experiment, two-dimensional electrophoresis was used to analyze the differential proteomics of the brain of 10- and 20-day-old suckling mice infected with attenuated rabies virus strain SRV9. The results showed that the expression levels of 10 protein spots had been up- or down-regulated at least 2-fold. Using MALDI-TOF-MS, we identified 8 differentially expressed proteins. We have identified proteins, namely hnRNP L, DPYSL3, NECAPs, and transaldolase that might be closely related to the susceptibility of SRV9 in suckling mice. Keywords: rabies virus; attenuated strain; suckling mouse; two-dimensional electrophoresis; proteomics.

Anatomical and physiological foundations of cerebello-hippocampal interaction.

Multiple lines of evidence suggest that functionally intact cerebello-hippocampal interactions are required for appropriate spatial processing. However, how the cerebellum anatomically and physiologically engages with the hippocampus to sustain such communication remains unknown. Using rabies virus as a retrograde transneuronal tracer in mice, we reveal that the dorsal hippocampus receives input from topographically restricted and disparate regions of the cerebellum. By simultaneously recording local field potential from both the dorsal hippocampus and anatomically connected cerebellar regions, we additionally suggest that the two structures interact, in a behaviorally dynamic manner, through subregion-specific synchronization of neuronal oscillations in the 6-12 Hz frequency range. Together, these results reveal a novel neural network macro-architecture through which we can understand how a brain region classically associated with motor control, the cerebellum, may influence hippocampal neuronal activity and related functions, such as spatial navigation.

Fundamentos teóricos del desempeño de los médicos y enfermeros en el manejo de la Rabia / Theoretical foundations of the doctors and nurses performance in the management of Rabies

Fundamento en el informe del año 2016, la Organización Mundial de la Salud reportó que 55 000 personas fallecen por rabia en África y Asia y la vacuna no es la mejor manera de prevenir la enfermedad por su elevado costo, en este análisis realizado por la Organización Mundial de la Salud, declara que el 95 porciento de los pacientes son menores de 15 años.Objetivo: describir la sistematización de los fundamentos teóricos que sustentan la preparación para el desempeño de médico y enfermero de la familia en el manejo de la rabia. Métodos: se realizó un estudio descriptivo en el período 2012 a noviembre de 2017, la investigación se sustenta desde la concepción dialéctico-materialista, para la obtención de los datos y el procesamiento de la información se emplearon como métodos teóricos (análisis documental, sistematización e histórico-lógico), médicos y enfermeros de la familia del municipio Boyeros.Resultados: la sistematización realizada permitió encontrar regularidades que se convierten en antecedentes a la definición operativa que se propone sobre desempeño del médico y enfermero de la familia en el manejo de la rabia a partir de los fundamentos de la educación médica como teoría educativa. Conclusiones: la sistematización realizada sobre el mejoramiento del desempeño del médico y enfermero de la familia, permitió establecer un acercamiento centrado en los referentes teóricos que sustentan el mejoramiento del desempeño del médico y enfermero de la familia, al identificar los principios de la Educación en el Trabajo para el desempeño de este personal en el manejo de la rabia(AU)

Background in the report of 2016, the World Health Organization (WHO) reported that 55 000 people died of rabies in Africa and Asia and the vaccine is not the best way to prevent the illness because of the high cost, in this analysis carried out by the WHO, declares 95 percent of patients are younger than 15 years old. Objective to describe the systematization the theoretical foundations that support the preparation for exert of doctor and nurse of family in the management of the Rabies. Methods: it was conducted a descriptive study in the period 2012-2017, the investigation is supported from the dialectic-materialist conception, for the obtaining of data and the processing the information was employed as theoretical methods (documentary analysis, systematization and historical-logical), doctors and nurses of family in the municipality Boyeros.Results: the carried out systematization allowed to find regularities that turn into antecedents for operative definition that proposes on exert of the doctor and nurse of family in the management of rabies from foundations of the medical education as educational theory. Conclusions: the carried out systematization on the improvement of the exert of the doctor and nurse of family, allowed to establish an approach centered in theoretical referents that support the improvement of exert of the doctor and nurse of family, identifying the principles of education in the work for the exert of these personnel in the management of rabies(AU)

Metabotropic glutamate receptor subtype 2 is a cellular receptor for rabies virus.

Rabies virus (RABV) invades the central nervous system and nearly always causes fatal disease in humans. How RABV interacts with host neuron membrane receptors to become internalized and cause rabid symptoms is not yet fully understood. Here, we identified a novel receptor of RABV, which RABV uses to infect neurons. We found that metabotropic glutamate receptor subtype 2 (mGluR2), a member of the G protein-coupled receptor family that is abundant in the central nervous system, directly interacts with RABV glycoprotein to mediate virus entry. RABV infection was drastically decreased after mGluR2 siRNA knock-down in cells. Antibodies to mGluR2 blocked RABV infection in cells in vitro. Moreover, mGluR2 ectodomain soluble protein neutralized the infectivity of RABV cell-adapted strains and a street strain in cells (in vitro) and in mice (in vivo). We further found that RABV and mGluR2 are internalized into cells and transported to early and late endosomes together. These results suggest that mGluR2 is a functional cellular entry receptor for RABV. Our findings may open a door to explore and understand the neuropathogenesis of rabies.

Rabies: changing prophylaxis and new insights in pathophysiology.

PURPOSE OF REVIEW: Despite great progress in decoding disease mechanisms, rabies remains one of the leading causes of human death worldwide. Towards the elimination of human rabies deaths by 2030, feasible and affordable post (PEP) and pre-exposure prophylaxis (PrEP) must be available with expansion to rural areas in rabies endemic countries. Vaccination and population control of dogs, principal reservoirs and transmitters, must be done in concert. RECENT FINDING: Advances in the understanding of rabies neuropathogenesis and pathophysiology are reviewed, including recent experimental findings on host- and virus-specific mechanisms mediating neuronal survival and explaining clinical differences in furious and paralytic rabies. The forthcoming World Health Organization guide on rabies based on pathogenesis and immunization mechanisms data with support by clinical evidence provide new accelerated 1 week intradermal PrEP and PEP schedules. Rabies immunoglobulin injected into the wound only is endorsed at amounts not exceeding the dose interfering with active immunization. Potential therapeutics as designed in accord with rabies neuro-pathophysiology are plausible. SUMMARY: Clinical practice and rabies awareness can be leveraged by transboundary collaboration among different areas. Advancement in prophylaxis and perspectives on animal control offer a new path to conquer rabies by 2030.

Comparative pathogenesis of rabies in bats and carnivores, and implications for spillover to humans.

Bat-acquired rabies is becoming increasingly common, and its diagnosis could be missed partly because its clinical presentation differs from that of dog-acquired rabies. We reviewed the scientific literature to compare the pathogenesis of rabies in bats and carnivores-including dogs-and related this pathogenesis to differences in the clinical presentation of bat-acquired and dog-acquired rabies in human beings. For bat-acquired rabies, we found that the histological site of exposure is usually limited to the skin, the anatomical site of exposure is more commonly the face, and the virus might be more adapted for entry via the skin than for dog-acquired rabies. These factors could help to explain several differences in clinical presentation between individuals with bat-acquired and those with dog-acquired rabies. A better understanding of these differences should improve the recording of a patient's history, enable drawing up of a more sophisticated list of clinical characteristics, and therefore obtain an earlier diagnosis of rabies after contact with a bat or carnivore that has rabies.

Rabies.

Rabies is a life-threatening neglected tropical disease: tens of thousands of cases are reported annually in endemic countries (mainly in Africa and Asia), although the actual numbers are most likely underestimated. Rabies is a zoonotic disease that is caused by infection with viruses of the Lyssavirus genus, which are transmitted via the saliva of an infected animal. Dogs are the most important reservoir for rabies viruses, and dog bites account for >99% of human cases. The virus first infects peripheral motor neurons, and symptoms occur after the virus reaches the central nervous system. Once clinical disease develops, it is almost certainly fatal. Primary prevention involves dog vaccination campaigns to reduce the virus reservoir. If exposure occurs, timely post-exposure prophylaxis can prevent the progression to clinical disease and involves appropriate wound care, the administration of rabies immunoglobulin and vaccination. A multifaceted approach for human rabies eradication that involves government support, disease awareness, vaccination of at-risk human populations and, most importantly, dog rabies control is necessary to achieve the WHO goal of reducing the number of cases of dog-mediated human rabies to zero by 2030.

Rabies - epidemiology, pathogenesis, public health concerns and advances in diagnosis and control: a comprehensive review.

Rabies is a zoonotic, fatal and progressive neurological infection caused by rabies virus of the genus Lyssavirus and family Rhabdoviridae. It affects all warm-blooded animals and the disease is prevalent throughout the world and endemic in many countries except in Islands like Australia and Antarctica. Over 60,000 peoples die every year due to rabies, while approximately 15 million people receive rabies post-exposure prophylaxis (PEP) annually. Bite of rabid animals and saliva of infected host are mainly responsible for transmission and wildlife like raccoons, skunks, bats and foxes are main reservoirs for rabies. The incubation period is highly variable from 2 weeks to 6 years (avg. 2-3 months). Though severe neurologic signs and fatal outcome, neuropathological lesions are relatively mild. Rabies virus exploits various mechanisms to evade the host immune responses. Being a major zoonosis, precise and rapid diagnosis is important for early treatment and effective prevention and control measures. Traditional rapid Seller's staining and histopathological methods are still in use for diagnosis of rabies. Direct immunofluoroscent test (dFAT) is gold standard test and most commonly recommended for diagnosis of rabies in fresh brain tissues of dogs by both OIE and WHO. Mouse inoculation test (MIT) and polymerase chain reaction (PCR) are superior and used for routine diagnosis. Vaccination with live attenuated or inactivated viruses, DNA and recombinant vaccines can be done in endemic areas. This review describes in detail about epidemiology, transmission, pathogenesis, advances in diagnosis, vaccination and therapeutic approaches along with appropriate prevention and control strategies.

Physical and chemical quality, biodiversity, and thermodynamic prediction of adhesion of bacterial isolates from a water purification system: a case study

ABSTRACT The objective of this study was to evaluate the quality of water purification system and identify the bacteria this system, predict bacterial adherence according to the hydrophobicity of these microorganisms and of the polypropylene distribution loop for purified water. The assessment of drinking water that supplies the purification system allowed good-quality physical, chemical, and microbiological specifications. The physicochemical specifications of the distributed purified water were approved, but the heterotrophic bacteria count was higher than allowed (>2 log CFU mL-1).The sanitation of the storage tank with chlorine decreased the number of bacteria adhered to the surface (4.34 cycles log). By sequencing of the 16SrDNA genes, six species of bacteria were identified. The contact angle was determined and polypropylene surface and all bacteria were considered to be hydrophilic, and adhesion was thermodynamically unfavorable. This case study showed the importance of monitoring the water quality in the purified water systems and the importance of sanitization with chemical agents. The count of heterotrophic bacteria on the polypropylene surface was consistent with the predicted thermodynamics results because the number of adhered cells reached approximate values of 5 log CFU cm-2.

Dogs that develop rabies post-vaccination usually manifest the paralytic subtype.

Rabies infection can manifest as either encephalitic (furious) or paralytic (dumb) types, with a ratio of approximately 2:1 in dogs. The clinical type of rabies that develops post-vaccination has only been reported in studies from one country, all with similar findings. We report a study of 36 rabid dogs with obtainable vaccination history, presenting to The Queen Saovabha Memorial Institute, Bangkok, Thailand during 2002-2008. Dogs were classified into encephalitic or paralytic types. Of 22 non-vaccinated dogs, 16 (73%) had the encephalitic type. In contrast, of the 14 vaccinated dogs, 10 (71%) had the paralytic type, a difference that was significant (p=0.016). Recent studies on canine brains have shown that lymphocyte response is more pronounced in paralytic rabies at the brainstem level, whereas viral burden is greater in the encephalitic form. We postulate partial immune response in the vaccinated dogs might influence rabies to manifest as the paralytic type. These results can serve as a natural experiment that can help explain the basis for the differences between the paralytic and encephalitic forms of canine rabies.

Quantitative Proteome Profiling of Street Rabies Virus-Infected Mouse Hippocampal Synaptosomes.

It is well established now that neuronal dysfunction rather than structural damage may be responsible for the development of rabies. In order to explore the underlying mechanisms in rabies virus (RABV) and synaptic dysfunctions, a quantitative proteome profiling was carried out on synaptosome samples from mice hippocampus. Synaptosome samples from mice hippocampus were isolated and confirmed by Western blot and transmission electron microscopy. Synaptosome protein content changes were quantitatively detected by Nano-LC-MS/MS. Protein functions were classified by the Gene Ontology (GO) and KEGG pathway. PSICQUIC was used to create a network. MCODE algorithm was applied to obtain subnetworks. Of these protein changes, 45 were upregulated and 14 were downregulated following RABV infection relative to non-infected (mock) synaptosomes. 28 proteins were unique to mock treatment and 12 were unique to RABV treatment. Proteins related to metabolism and synaptic vesicle showed the most changes in expression levels. Furthermore, protein-protein interaction (PPI) networks revealed that several key biological processes related to synaptic functions potentially were modulated by RABV, including energy metabolism, cytoskeleton organization, and synaptic transmission. These data will be useful for better understanding of neuronal dysfunction of rabies and provide the foundation for future research.

Household exposure and animal-bite surveillance following human rabies detection in Southern Ghana.

INTRODUCTION: Rabies remains a neglected tropical zoonotic disease with 100% case fatality rate and estimated 6,000 global mortality annually, and yet vaccine preventable. In Ghana, rabies outbreaks receive poor response. We investigated rabies in a 5-year old boy to find the source of infection, identify exposed persons for post-exposure prophylaxis and describe animal-bite surveillance in Manya-Krobo District of Ghana. METHODS: We actively searched for cases and exposures by interviewing household members of the victim, schoolmates, and health professionals using WHO case definition, interview guide and checklist. We reviewed health and veterinary records and reports, and interviewed stakeholders. Descriptive data analyses were carried out and presented using tables and charts. Recorded responses were transcribed into thematic areas and analysed. RESULTS: Child had dog-bite at the wrist, and developed hyperactivity, hydrophobia and hyperventilation 2 months post bite. He was hospitalised and died from respiratory failure day 3 after admission. Thirty-three persons were exposed to rabies infectious material. Females were 66%, age-groups 5-15yrs and 30-59 yrs were 33.3% and 39.4% respectively. A third (11/33) were category II exposure by WHO classification and were recommended for post-exposure prophylaxis. Surveillance records showed ninety-two animal-bite cases were reported for past 12 months. Half were females, and 18-59yrs age-group was 43%. Surveillance data quality was poor. CONCLUSION: Rabies remains a public health burden inGhana with domestic dog as reservoir of the virus and females more vulnerable to secondary exposures. Health education on rabies should be intensified, and robust animal-bite surveillance system put in place.

Diabolical effects of rabies encephalitis.

Rabies is an acute encephalomyelitis in humans and animals caused by rabies virus (RABV) infection. Because the neuropathological changes are very mild in rabies, it has been assumed that neuronal dysfunction likely explains the severe clinical disease. Recently, degenerative changes have been observed in neuronal processes (dendrites and axons) in experimental rabies. In vitro studies have shown evidence of oxidative stress that is caused by mitochondrial dysfunction. Recent work has shown that the RABV phosphoprotein (P) interacts with mitochondrial Complex I leading to overproduction of reactive oxygen species, which results in injury to axons. Amino acids at positions 139 to 172 of the P are critical in this process. Rabies vectors frequently show behavioral changes. Aggressive behavior with biting is important for transmission of the virus to new hosts at a time when virus is secreted in the saliva. Aggression is associated with low serotonergic activity in the brain. Charlton and coworkers performed studies in experimentally infected striped skunks with skunk rabies virus and observed aggressive behavioral responses. Heavy accumulation of RABV antigen was found in the midbrain raphe nuclei, indicating that impaired serotonin neurotransmission from the brainstem may account for the aggressive behavior. We now have an improved understanding of how RABV causes neuronal injury and how the infection results in behavioral changes that promote viral transmission to new hosts.

Viruses, apoptosis, and neuroinflammation--a double-edged sword.

Apoptosis, or programmed cell death, is a fundamental and widespread cell biological process that is distinct from cell necrosis and can be induced by a wide variety of stimuli including viral infections. Apoptosis may occur via either the intrinsic or extrinsic pathways and confers several advantages to the virally infected host including the prevention of further viral propagation and the potential inhibition and resolution of inflammatory processes. Several viruses have been shown to have the capacity to induce apoptosis in susceptible cells including herpes simplex virus, Varicella-zoster virus, rabies virus, human immunodeficiency virus, and reovirus. Apoptosis has also been observed in human African trypanosomiasis which is an infection caused by a protozoan parasite. The mechanisms leading to apoptosis may differ depending on the type of infection. Apoptosis has been reported in several neurodegenerative diseases and also psychiatric disorders but the true clinical significance of such observations is not certain, and, though interesting, it is very difficult to ascribe causation in these conditions. The presence of inflammation in the central nervous system in any neurological condition, including those associated with a viral infection, is not necessarily an absolute marker of serious disease and the notion of 'good' versus 'bad' inflammation is considered to be valid in some circumstances. The precise relationship between viruses, apoptosis, and inflammation is viewed as a complex one requiring further investigation to unravel and understand its nature.