Tracey McNamara
Updates on West Nile Virus
It has been 13 years since West Nile virus (WNV) first appeared in North America. Although some progress has been made in the understanding of its neurotropism, pathogenicity, modes of transmission, and species susceptibility, many unanswered questions remain about this resourceful and devastating virus. It is not hard to see how much has changed by just looking back at what was believed in 1999. At that time, it was thought that WNV could only be spread through the bite of a mosquito; that it usually caused a mild febrile and self-limiting infection; that only the very young and very old were at risk of developing neuroinvasive disease that affected only the brain; that mammals were dead-end hosts. By 2002, everything that had been said about WNV had to be reevaluated. By then, it had been shown that WNV could, in fact, be spread by many routes other than mosquito bite;6,7 that even mild infections might not be benign and may cause long-term sequelae such as fatigue and cognitive deficits;5 that other risk factors might be involved in development of West Nile neuroinvasive disease (WNND); that patients with WNND could also present with poliomyelitis18 and serious long-term deficits such as Parkinsonism-like disorders;23 that mammals are not necessarily dead-end hosts and may play a role in the epidemiology of WNV spread; and so on. And we are still learning.
Steele et al.58 described WNV infection and illness in a number of avian zoo species in 2000 and suggested that species other than crows were susceptible. Although corvids are certainly excellent sentinels for WNV, the list of known susceptible avian species is now lengthy.29 We also now know that species as diverse as alligators,36 polar bears,16 reindeer,43 harbor seals and gray seals,15 killer whales,57 Barbary macaques,41 and psittacines44 may succumb to WNV infection. Ducks, which had been believed to be resistant to WNV infection, have also experienced die-offs.22 Unusual cutaneous lesions were described in American alligators.36 Species other than crows such as blue jays and sparrows17,39 have emerged as good indicators of WNV activity. Mammals such as fox squirrels (Sciurus niger),51 eastern chipmunks (Tamias striatus),48 and eastern cottontail rabbits (Sylvagus floridanus)25 have been found to develop viremias sufficient to infect mosquitoes and may play a role in the epidemiologic spread of WNV in urban environments. WNV infection has also had significant impacts on raptors. Clinical and pathologic findings have been described elsewhere.32 Raptors also emerged as an excellent surveillance tool. In one study, raptor admissions to rehabilitation clinics took place 14 weeks earlier than other surveillance methods.34,35 Ophthalmologic lesions were described both in humans26 and birds,46 and this led to the development of a rapid, safe sampling method in crows called the “intraocular cocktail” that involves inserting a needle through the cornea and vigorously scraping the intraocular contents, aspirating them, and processing the material for nucleic acid extraction.30 Feather testing has also proven useful.35 Mosquito saliva was found to enhance WNV infection in mice bitten by Culex tarsalis, which suggests that it exerts a local effect.62
Steele et al.58 also demonstrated abundant WNV antigens in the kidneys and intestinal tracts of infected zoo birds and suggested that direct horizontal transmission might be taking place among birds. Demonstration of antigen in ovarian and testicular tissue also hinted at the possibility of vertical transmission. We now know the virus may be transmitted directly bird to bird.31 Studies have demonstrated oral and cloacal shedding of WNV,28 which ultimately led to the development of a rapid diagnostic test used by many zoos and health departments across the nation.45,59,60 WNV may also be spread via oral transmission,2,53 via breast milk,2 via the intrauterine route,2 and via organ transplantation9 and blood transfusions. Perhaps the fact that, as shown by experimental studies in primates,49 hamsters,70 and mice,1 WNV may persist for months in infected animals is of greatest concern. Most recently, human studies have confirmed that viral persistence is present years after infection and is associated with the development of chronic kidney disease.37 What this means for humans or animals has yet to be elucidated, but viral persistence may be associated with well-documented long-term sequelae seen in people.
The majority of recent literature has focused on the molecular biology of WNV. This chapter will summarize some of the recent work in that area, as well as information on viral persistence, long-term sequelae, and recent vaccine efforts.
In the past decade, much has been learned about the structure of the WNV virion. The WNV virion has several key structural proteins: the capsid protein C that binds viral ribonucleic acid (RNA), the premembrane (prM) protein that blocks premature viral fusion; and an E protein that mediates viral attachment, membrane fusion, and viral assembly. The majority of neutralizing antibodies are directed against regions of the E protein, although a subset likely recognizes prM.52 The virion also has nonstructural proteins that regulate viral transcription and replication and attenuate host antiviral responses.52 In 2002, a new strain of WNV emerged, but it differed from the NY99 strain in the envelope (E) protein at amino acid 159 (WNV02).13 This is now the dominant genotype in North America. This single amino acid change has led to increased intensity of transmission of WNV and rapid geographic expansion.13
How WNV crosses the blood–brain barrier (BBB) and whether central nervous system (CNS) damage is caused by direct viral infection, indirectly by the host’s immune response, or both, is a question that has been the subject of many studies.8,10,13,19,20,33,52,54–56,66,67 Entry into the CNS is most likely through hematogenous spread with the help of tumor necrosis factor–alpha (TNF-α) and matrix metalloproteinases (MMPs), which increase the permeability of the BBB.52,66 Adhesion molecules on the vascular endothelium and leukocytes play important roles in controlling entry into the CNS. Intercellular adhesion molecule 1 (ICAM-1) is critical to this process and plays an important role in neuroinvasion in mice.10 Leukocyte trafficking to the CNS has been linked to the chemokine receptor CCR5, which is upregulated by WNV infection and is associated with CNS infiltration of CD+4 and CD+8 T cells, natural killer cells, and macrophages expressing the receptor.19 CD+8 cells control infection by producing antiviral cytokines (interferon [IFN] or TNF-α) early in infection or by triggering the death of WNV-infected cells through perforin or FAS ligand–dependent pathways.56 TNF-related apoptosis-inducing ligand (TRAIL) produced by CD+8 T-cells contributes to disease resolution by helping to clear WNV from the neurons in the CNS.55 However, WNV has developed strategies for enhancing viral replication in the host by blocking the action of type 1-IFN and evading the antiviral activity of IFN-stimulated genes.33 Two human genes, CCR5 and 2′5′ oligoadenylate synthetase (OAS1b), have been identified as susceptible loci for WNV infection.
Long-Term Sequelae
In 1999, WNV was believed to cause a mild, febrile, self-limiting illness in the majority of patients, with a small percentage developing neuroinvasive disease. By 2004, it was recognized that WNV infection could result in a protracted convalescent period with long-term problems with memory; confusion; clinical depression; muscle weakness; tremors; and parkinsonism-like disorders21,27 18 months after infection. In a Houston study, 60% of encephalitic patients reported signs 5 years after infection.65 Patients with a milder form of illness are just as likely to suffer long-term health problems as encephalitis cases.4 In this study of WN fever, 84% of patients reported persistent fatigue, 59% memory problems, and 49% ongoing muscle weakness. Studies on long-term sequelae in naturally infected animals are not available. It is postulated that long-term sequelae may be a result of viral persistence.
Viral Persistence
In 1983, Pogodina et al.49 published a study on WNV persistence in primates experimentally infected with several strains of WNV. They found that encephalitis was present in animals with neuroinvasive disease, with only febrile illness, or with asymptomatic infections. This was considered unusual, as most RNA viral infections are transient and are subsequently cleared by the host.1 Concerned about this possibility, the Department of Pathology at the Bronx Zoo began long-term studies on known positive WNV cases in the zoo collection. In 2000, evaluation of brain tissue from known positive animals suggested that viral persistence might be taking place with the NY99 strain of WNV in naturally infected animals. A symptomatic snow leopard (Panthera uncia) and a greater Indian rhinoceros (Rhinoceros unicornis) died 3 and 8 months following illness and seroconversion. Both exhibited dramatic lymphoplasmacytic cuffing in the CNS at the time of death. An asymptomatic but seropositive babirousa (Babyrousa babyrussa) that died 10 months after seroconversion also had mild to moderate lymphoplasmacytic cuffing in the brain, which suggested that subclinical infections may also produce CNS pathology.
In 2006, WNV was demonstrated in the brain and CSF of an immunocompromised patient 4 months after initial diagnosis and in spite of treatment with IFN, immunoglobulins, and ribavirin. Persistent infection with WNV was demonstrated by the identification of WNV nucleic acid in the brain by reverse-transcriptase polymerase chain reaction (RT-PCR) assay and immunohistochemical demonstration of antigen in tissue.47 A 2010 study1 developed a murine model of viral persistence. It found viral persistence in the face of a robust antibody response and in the presence of inflammation in the brain even in subclinical infections and showed that WNV persisted in the CNS and peripheral tissues for up to 6 months following infection in mice with subclinical infections. The authors concluded that the “frequency, duration, and tissue location of WNV persistence are species dependent, probably due to differences in host immunity, severity of disease, initial viral loads and tissue tropism, and cell targets.”1 These studies have raised concerns about the role of viral persistence in the development of the now-recognized long-term sequelae of WNV infection. They also raise concerns about the estimated 1.2 million people with asymptomatic WNV infections in the United States and possible subclinical CNS disease.
The kidney has also been a focus of persistence studies. In 2005, WNV was detected in the urine of a patient with encephalitis 8 days after symptom onset. Viral RNA was detected by RT-PCR.64 In another 2005 study in hamsters,63 chronic renal infection and persistent shedding was found in urine up to 8 months after infection. When the isolates that resulted in renal tropism were compared with the wild-type parent virus (NY 385-99), nucleotide changes were found in coding regions, causing amino acid substitutions in the E, NS1, NS2B, and NS5 proteins.14 A 2012 long-term study of patients in Houston found an association between neuroinvasive WNV infection and the development of chronic kidney disease and suggested that all patients with WNV should have their renal function closely monitored.33
It is not known what impact viral persistence may have on zoo species, but many zoo species tested WNV positive on plaque reduction neutralization test (PRNT) or RT-PCR during the national zoological surveillance for WNV project and, like humans, the animals also should have long-term follow-up. Unlike the human medical community, zoo practitioners have ready access to necropsy material for study. The zoo community is in the position to render a great service to public health, given its ability to follow animals over the long term and evaluate them for viral persistence and renal and neuropathology. Tissues should be harvested and frozen at −80° C for virus isolation and RT-PCR from all known WNV-positive birds and mammals at the time of necropsy. Any zoo clinician or pathologist who performs a necropsy on a previously positive animal, especially a long-lived mammal, is urged to contact this author. A grant is in preparation to cover the costs of virus isolation (VI), RT-PCR, histopathology, and immunohistochemistry on WNV survivors.
Stay updated, free articles. Join our Telegram channel
Full access? Get Clinical Tree
