Outbreak Of Human Encephalitis Research Articles

Shifts in receptors during submergence of an encephalitic arbovirus

Western equine encephalitis virus (WEEV) is an arthropod-borne virus (arbovirus) that frequently caused major outbreaks of encephalitis in humans and horses in the early twentieth century, but the frequency of outbreaks has since decreased markedly, and strains of this alphavirus isolated in the past two decades are less virulent in mammals than strains isolated in the 1930s and 1940s1–3. The basis for this phenotypic change in WEEV strains and coincident decrease in epizootic activity (known as viral submergence3) is unclear, as is the possibility of re-emergence of highly virulent strains. Here we identify protocadherin 10 (PCDH10) as a cellular receptor for WEEV. We show that multiple highly virulent ancestral WEEV strains isolated in the 1930s and 1940s, in addition to binding human PCDH10, could also bind very low-density lipoprotein receptor (VLDLR) and apolipoprotein E receptor 2 (ApoER2), which are recognized by another encephalitic alphavirus as receptors4. However, whereas most of the WEEV strains that we examined bind to PCDH10, a contemporary strain has lost the ability to recognize mammalian PCDH10 while retaining the ability to bind avian receptors, suggesting WEEV adaptation to a main reservoir host during enzootic circulation. PCDH10 supports WEEV E2–E1 glycoprotein-mediated infection of primary mouse cortical neurons, and administration of a soluble form of PCDH10 protects mice from lethal WEEV challenge. Our results have implications for the development of medical countermeasures and for risk assessment for re-emerging WEEV strains.

A Conserved Basic Patch and Central Kink in the Nipah Virus Phosphoprotein Multimerization Domain Are Essential for Polymerase Function

Nipah virus is a highly lethal zoonotic pathogen found in Southeast Asia that has caused human encephalitis outbreaks with 40%-70% mortality. NiV encodes its own RNA-dependent RNA polymerase within the large protein, L. Efficient polymerase activity requires the phosphoprotein, P, which tethers L to its template, the viral nucleocapsid. P is a multifunctional protein with modular domains. The central P multimerization domain is composed of a long, tetrameric coiled coil. We investigated the importance of structural features found in this domain for polymerase function using a newly constructed NiV bicistronic minigenome assay. We identified a conserved basic patch and central kink in the coiled coil that are important for polymerase function, with R555 being absolutely essential. This basic patch and central kink are conserved in the related human pathogens measles and mumps viruses, suggesting that this mechanism may be conserved.

CCR2 Plays a Protective Role in Rocio Virus-Induced Encephalitis by Promoting Macrophage Infiltration Into the Brain.

Rocio virus (ROCV) is a highly neuropathogenic mosquito-transmitted flavivirus responsible for an unprecedented outbreak of human encephalitis during 1975–1976 in Sao Paulo State, Brazil. Previous studies have shown an increased number of inflammatory macrophages in the central nervous system (CNS) of ROCV-infected mice, implying a role for macrophages in the pathogenesis of ROCV. Here, we show that ROCV infection results in increased expression of CCL2 in the blood and in infiltration of macrophages into the brain. Moreover, we show, using CCR2 knockout mice, that CCR2 expression is essential for macrophage infiltration in the brain during ROCV infection and that the lack of CCR2 results in increased disease severity and mortality. Thus, our findings show the protective role of CCR2-mediated infiltration of macrophages in the brain during ROCV infection.

West Nile and St. Louis encephalitis viral genetic determinants of avian host competence.

West Nile virus (WNV) and St. Louis encephalitis (SLEV) virus are enzootically maintained in North America in cycles involving the same mosquito vectors and similar avian hosts. However, these viruses exhibit dissimilar viremia and virulence phenotypes in birds: WNV is associated with high magnitude viremias that can result in mortality in certain species such as American crows (AMCRs, Corvus brachyrhynchos) whereas SLEV infection yields lower viremias that have not been associated with avian mortality. Cross-neutralization of these viruses in avian sera has been proposed to explain the reduced circulation of SLEV since the introduction of WNV in North America; however, in 2015, both viruses were the etiologic agents of concurrent human encephalitis outbreaks in Arizona, indicating the need to re-evaluate host factors and cross-neutralization responses as factors potentially affecting viral co-circulation. Reciprocal chimeric WNV and SLEV viruses were constructed by interchanging the pre-membrane (prM)-envelope (E) genes, and viruses subsequently generated were utilized herein for the inoculation of three different avian species: house sparrows (HOSPs; Passer domesticus), house finches (Haemorhous mexicanus) and AMCRs. Cross-protective immunity between parental and chimeric viruses were also assessed in HOSPs. Results indicated that the prM-E genes did not modulate avian replication or virulence differences between WNV and SLEV in any of the three avian species. However, WNV-prME proteins did dictate cross-protective immunity between these antigenically heterologous viruses. Our data provides further evidence of the important role that the WNV / SLEV viral non-structural genetic elements play in viral replication, avian host competence and virulence.

Helicase Domain of West Nile Virus NS3 Protein Plays a Role in Inhibition of Type I Interferon Signalling.

West Nile virus (WNV) is a neurotropic flavivirus that can cause encephalitis in mammalian and avian hosts. In America, the virulent WNV strain (NY99) is causing yearly outbreaks of encephalitis in humans and horses, while in Australia the less virulent Kunjin strain of WNV strain has not been associated with significant disease outbreaks until a recent 2011 large outbreak in horses (but not in humans) caused by NSW2011 strain. Using chimeric viruses between NY99 and NSW2011 strains we previously identified a role for the non-structural proteins of NY99 strain and especially the NS3 protein, in enhanced virus replication in type I interferon response-competent cells and increased virulence in mice. To further define the role of NY99 NS3 protein in inhibition of type I interferon response, we have generated and characterised additional chimeric viruses containing the protease or the helicase domains of NY99 NS3 on the background of the NSW2011 strain. The results identified the role for the helicase but not the protease domain of NS3 protein in the inhibition of type I interferon signalling and showed that helicase domain of the more virulent NY99 strain performs this function more efficiently than helicase domain of the less virulent NSW2011 strain. Further analysis with individual amino acid mutants identified two amino acid residues in the helicase domain primarily responsible for this difference. Using chimeric replicons, we also showed that the inhibition of type I interferon (IFN) signalling was independent of other known functions of NS3 in RNA replication and assembly of virus particles.

Estimating biologically relevant parameters under uncertainty for experimental within-host murine West Nile virus infection.

West Nile virus (WNV) is an emerging pathogen that has decimated bird populations and caused severe outbreaks of viral encephalitis in humans. Currently, little is known about the within-host viral kinetics of WNV during infection. We developed mathematical models to describe viral replication, spread and host immune response in wild-type and immunocompromised mice. Our approach fits a target cell-limited model to viremia data from immunocompromised knockout mice and an adaptive immune response model to data from wild-type mice. Using this approach, we first estimate parameters governing viral production and viral spread in the host using simple models without immune responses. We then use these parameters in a more complex immune response model to characterize the dynamics of the humoral immune response. Despite substantial uncertainty in input parameters, our analysis generates relatively precise estimates of important viral characteristics that are composed of nonlinear combinations of model parameters: we estimate the mean within-host basic reproductive number,R0, to be 2.3 (95% of values in the range 1.7-2.9); the mean infectious virion burst size to be 2.9 plaque-forming units (95% of values in the range 1.7-4.7); and the average number of cells infected per infectious virion to be between 0.3 and 0.99. Our analysis gives mechanistic insights into the dynamics of WNV infection and produces estimates of viral characteristics that are difficult to measure experimentally. These models are a first step towards a quantitative understanding of the timing and effectiveness of the humoral immune response in reducing host viremia and consequently the epidemic spread of WNV.

Evaluation of vector competence for West Nile virus in Italian Stegomyia albopicta (=Aedes albopictus) mosquitoes.

West Nile virus (WNV) is a zoonotic arboviral pathogen transmitted by mosquitoes in a cycle that involves wild birds as reservoir hosts. The virus is responsible for outbreaks of viral encephalitis in humans and horses. In Europe, Culex pipiens (Diptera: Culicidae) is considered to be the main vector of WNV, but other species such as Stegomyia albopicta (=Aedes albopictus) (Diptera: Culicidae) may also act as competent vectors of this virus. Since 2008 human cases of WNV disease have been reported in northeast Italy. In 2011, new areas of southern Italy became involved and a first outbreak of WNV lineage 1 occurred on the island of Sardinia. On the assumption that a potential involvement of St. albopicta in WNV transmission cannot be excluded, and in order to evaluate the competence of this species for the virus, an experimental infection of an St. albopicta laboratory colony, established from mosquitoes collected in Sardinia, was carried out. The results were compared with those obtained in a colony of the main vector Cx. pipiens. The study showed St. albopicta collected on Sardinia to be susceptible to WNV infection, which suggests this Italian mosquito species is able to act as a possible secondary vector, particularly in urban areas where the species reaches high levels of seasonal abundance.

The Australian Public is Still Vulnerable to Emerging Virulent Strains of West Nile Virus.

The mosquito-borne West Nile virus (WNV) is responsible for outbreaks of viral encephalitis in humans and horses with particularly virulent strains causing recent outbreaks in Eastern Europe, the Middle East, and North America. In Australia, a strain of WNV, Kunjin (WNVKUN), is endemic in the north and infection with this virus is generally asymptomatic. However, in early 2011, following extensive flooding, an unprecedented outbreak of WNVKUN encephalitis in horses occurred in South-Eastern Australia, resulting in more than 1,000 cases and a mortality of 10–15%. Despite widespread evidence of equine infections, there was only a single mild human case reported during this outbreak. To understand why clinical disease was seen in horses without similar observations in the human population, a serosurvey was conducted using blood donor samples from areas where equine cases were reported to assess level of flavivirus exposure. The seroprevalence to WNVKUN in humans was low before the outbreak (0.7%), and no significant increase was demonstrated after the outbreak period (0.6%). Due to unusual epidemiological features during this outbreak, a serosurvey was also conducted in rabbits, a potential reservoir host. Out of 675 animals, sampled across Australia between April 2011 and November 2012, 86 (12.7%) were seropositive for WNVKUN, with the highest prevalence during February of 2012 (28/145; 19.3%). As this is the first serological survey for WNVKUN in Australian feral rabbits, it remains to be determined whether wild rabbits are able to develop a high enough viremia to actively participate in WNV transmission in Australia. However, they may constitute a sentinel species for arbovirus activity, and this is the focus of on-going studies. Collectively, this study provides little evidence of human exposure to WNVKUN during the 2011 outbreak and indicates that the Australian population remains susceptible to the emergence of virulent strains of WNV.

The Changing Epidemiology of Kunjin Virus in Australia

West Nile virus (WNV) is a mosquito-borne virus responsible for outbreaks of viral encephalitis in humans and horses, with particularly virulent strains causing recent outbreaks of disease in Eastern Europe, the Middle East and North America. A strain of WNV, Kunjin (WNVKUN), is endemic in northern Australia and infection with this virus is generally asymptomatic. However in early 2011, an unprecedented outbreak of encephalitis in horses occurred in south-eastern Australia, resulting in mortality in approximately 10%–15% of infected horses. A WNV-like virus (WNVNSW2011) was isolated and found to be most closely related to the indigenous WNVKUN, rather than other exotic WNV strains. Furthermore, at least two amino acid changes associated with increased virulence of the North American New York 99 strain (WNVNY99) compared to the prototype WNVKUN were present in the WNVNSW2011 sequence. This review summarizes our current understanding of WNVKUN and how the epidemiology and ecology of this virus has changed. Analysis of virulence determinants of contemporary WNVKUN isolates will provide clues on where virulent strains have emerged in Australia. A better understanding of the changing ecology and epidemiology associated with the emergence of virulent strains is essential to prepare for future outbreaks of WNV disease in Australia.

Influence of the CCR-5/MIP-1 α Axis in the Pathogenesis of Rocio Virus Encephalitis in a Mouse Model

Rocio virus (ROCV) caused an outbreak of human encephalitis during the 1970s in Brazil and its immunopathogenesis remains poorly understood. CC-chemokine receptor 5 (CCR5) is a chemokine receptor that binds to macrophage inflammatory protein (MIP-1 α). Both molecules are associated with inflammatory cells migration during infections. In this study, we demonstrated the importance of the CCR5 and MIP-1 α, in the outcome of viral encephalitis of ROCV-infected mice. CCR5 and MIP-1 α knockout mice survived longer than wild-type (WT) ROCV-infected animals. In addition, knockout mice had reduced inflammation in the brain. Assessment of brain viral load showed mice virus detection five days post-infection in wild-type and CCR5-/- mice, while MIP-1 α-/- mice had lower viral loads seven days post-infection. Knockout mice required a higher lethal dose than wild-type mice as well. The CCR5/MIP-1 α axis may contribute to migration of infected cells to the brain and consequently affect the pathogenesis during ROCV infection.

Structure of the St. Louis Encephalitis Virus Postfusion Envelope Trimer

St. Louis encephalitis virus (SLEV) is a mosquito-borne flavivirus responsible for several human encephalitis outbreaks over the last 80 years. Mature flavivirus virions are coated with dimeric envelope (E) proteins that mediate attachment and fusion with host cells. E is a class II fusion protein, the hallmark of which is a distinct dimer-to-trimer rearrangement that occurs upon endosomal acidification and insertion of hydrophobic fusion peptides into the endosomal membrane. Herein, we report the crystal structure of SLEV E in the posfusion trimer conformation. The structure revealed specific features that differentiate SLEV E from trimers of related flavi- and alphaviruses. SLEV E fusion loops have distinct intermediate spacing such that they are positioned further apart than previously observed in flaviviruses but closer together than Semliki Forest virus, an alphavirus. Domains II and III (DII and DIII) of SLEV E also adopt different angles relative to DI, which suggests that the DI-DII joint may accommodate spheroidal motions. However, trimer interfaces are well conserved among flaviviruses, so it is likely the differences observed represent structural features specific to SLEV function. Analysis of surface potentials revealed a basic platform underneath flavivirus fusion loops that may interact with the anionic lipid head groups found in membranes. Taken together, these results highlight variations in E structure and assembly that may direct virus-specific interactions with host determinants to influence pathogenesis.

Virus del oeste del Nilo: perspectivas en el mundo vertebrado

El virus del Oeste del Nilo (VON) pertenece a la familia Flaviviridae, género Flavivirus, es transmitido por artrópodos en un ciclo que involucra a mosquitos y aves. Por muchas décadas había sido reconocido en África, Asia y el sur de Europa. El virus apareció por primera vez en Estados Unidos en año de 1999 y se ha documentado su circulación en México, Islas Caimán, Jamaica, República Dominicana, Martinica, Guadalupe, Cuba, Puerto Rico, El Salvador y Colombia. Sólo recientemente fue asociado con un creciente número de brotes de encefalitis en humanos y equinos, además de infecciones en vertebrados de una gran variedad de especies. Muchos animales, incluyendo más de 150 especies de aves y al menos 30 vertebrados de otras especies son susceptibles a la infección por VON. El resultado de las infecciones depende de la especie, la edad del animal, su estado inmune y la patogenicidad de la cepa del virus. La infección ocurre en aves, como passeriformes, pollos jóvenes y gansos domésticos, lo que resulta en altos títulos de viremia que permite la transmisión a los artrópodos. Está menos caracterizado el rol de los vertebrados en la transmisión indirecta, como por órganos contaminados, tejidos o excreciones. La creciente importancia de la infección por VON ha conducido al desarrollo de vacunas veterinarias con virus muertos, atenuados, quiméricos y vacunas de ADN recombinante. En esta revisión se discute en forma general la importancia de infección por VON en vertebrados, su rol en medicina veterinaria y su posible impacto en salud animal por su reciente introducción a Colombia.

West Nile virus in the vertebrate world

West Nile virus (WNV), an arthropod-borne virus belonging to the family Flaviviridae, had been recognized in Africa, Asia and the south of Europe for many decades. Only recently, it has been associated with an increasing number of outbreaks of encephalitis in humans and equines as well as an increasing number of infections in vertebrates of a wide variety of species. In this article, the data available on the incidence of WNV in vertebrates are reviewed. Moreover, the role of vertebrates in the transmission of WNV, the control of WNV infections in veterinary medicine as well as future perspectives are discussed. A wide variety of vertebrates, including more than 150 bird species and at least 30 other vertebrate species, are susceptible to WNV infection. The outcome of infection depends on the species, the age of the animal, its immune status and the pathogenicity of the WNV isolate. WNV infection of various birds, especially passeriforms, but also of young chickens and domestic geese, results in high-titred viremia that allows arthropod-borne transmission. For other vertebrate species, only lemurs, lake frogs and hamsters develop suitable viremia levels to support arthropod-borne transmission. The role of vertebrates in direct, non-arthropod-borne transmission, such as via virus-contaminated organs, tissues or excretions is less well characterized. Even though direct transmission can occur among vertebrates of several species, data are lacking on the exact amounts of infectious virus needed. Finally, the increased importance of WNV infections has led to the development of killed, live-attenuated, DNA-recombinant and chimeric veterinary vaccines.

Exposure of domestic mammals to West Nile virus during an outbreak of human encephalitis, New York City, 1999.

We evaluated West Nile (WN) virus seroprevalence in healthy horses, dogs, and cats in New York City after an outbreak of human WN virus encephalitis in 1999. Two (3%) of 73 horses, 10 (5%) of 189 dogs, and none of 12 cats tested positive for WN virus-neutralizing antibodies. Domestic mammals should be evaluated as sentinels for local WN virus activity and predictors of the infection in humans.

Exposure of Domestic Mammals to West Nile Virus during an Outbreak of Human Encephalitis, New York City, 1999

Abstract We evaluated West Nile (WN) virus seroprevalence in healthy horses, dogs, and cats in New York City after an outbreak of human WN virus encephalitis in 1999. Two (3%) of 73 horses, 10 (5%) of 189 dogs, and none of 12 cats tested positive for WN virus- neutralizing antibodies. Domestic mammals should be evaluated as sentinels for local WN virus activity and predictors of the infection in humans.

Detection of arboviral RNA directly from mosquito homogenates by reverse-transcription–polymerase chain reaction

Many arthropod-borne viruses (arboviruses) are important human pathogens medically. The development of an effective technique to detect the viruses by using nucleic acid amplification, such as polymerase chain reaction (PCR), improves not only clinical diagnosis but also virologic surveillance of mosquito vectors in the field. In this study, the development of an improved and simplified assay is described for detection of mosquitoes infected with eastern equine encephalitis (EEE) virus, Cache Valley (CV), and California (CAL) serogroup viruses from field-collected mosquito pools. As little as 5 μl of homogenate from mosquito pools was used in the reverse transcription (RT) reaction followed by the use of three sets of specific primers for the PCR. Positive pools were determined by finding PCR bands of the expected size for each arbovirus. The confirmation and identification of Bunyaviruses was done by sequencing the PCR product. In 1999, West Nile virus (WNV) was identified as the etiologic agent of an outbreak of human encephalitis in New York City. It is shown that this protocol is also able to detect West Nile viral RNA in a pool of 100 mosquitoes containing one infected mosquito.

Origin of the West Nile virus responsible for an outbreak of encephalitis in the northeastern United States.

In late summer 1999, an outbreak of human encephalitis occurred in the northeastern United States that was concurrent with extensive mortality in crows (Corvus species) as well as the deaths of several exotic birds at a zoological park in the same area. Complete genome sequencing of a flavivirus isolated from the brain of a dead Chilean flamingo (Phoenicopterus chilensis), together with partial sequence analysis of envelope glycoprotein (E-glycoprotein) genes amplified from several other species including mosquitoes and two fatal human cases, revealed that West Nile (WN) virus circulated in natural transmission cycles and was responsible for the human disease. Antigenic mapping with E-glycoprotein-specific monoclonal antibodies and E-glycoprotein phylogenetic analysis confirmed these viruses as WN. This North American WN virus was most closely related to a WN virus isolated from a dead goose in Israel in 1998.

Emergence of a new arbovirus disease in Brazil. II. Epidemiologic studies on 1975 epidemic.

In April, 1975, an outbreak of human encephalitis caused by Rocio virus, a new flavivirus, occurred in the coastal region of southern São Paulo, Brazil. The epidemic started during the second half of March, 1975, and ended in June, 1975. A total of 465 cases with 61 deaths were recorded, and the overall attack rate was 15 per thousand population. The mortality rate was two per thousand inhabitants and the overall case-fatality rate was 13%. However, the case-fatality rate was markedly decreased when adequate hospital care was provided. The number of cases in the 15--30 years age group was higher than in the other age groups and the rate was higher in males than in females. Lack of evidence of person-to-person transmission, based on a study of families involved in the epidemic and on the antigenic relationship to other mosquito-borne flaviviruses, suggested that the virus was transmitted by a vector and that wild animals were involved in the virus cycle.