Anna Grahn

Recombination of Globally Circulating Varicella Zoster Virus

ABSTRACT Varicella-zoster virus (VZV) is a human herpesvirus, which during primary infection typically causes varicella (chicken pox) and establishes lifelong latency in sensory and autonomic ganglia. Later in life, the virus may reactivate to cause herpes zoster (HZ; also known as shingles). To prevent these diseases, a live-attenuated heterogeneous vaccine preparation, vOka, is used routinely in many countries worldwide. Recent studies of another alphaherpesvirus, infectious laryngotracheitis virus, demonstrate that live-attenuated vaccine strains can recombine in vivo, creating virulent progeny. These findings raised concerns about using attenuated herpesvirus vaccines under conditions that favor recombination. To investigate whether VZV may undergo recombination, which is a prerequisite for VZV vaccination to create such conditions, we here analyzed 115 complete VZV genomes. Our results demonstrate that recombination occurs frequently for VZV. It thus seems that VZV is fully capable of recombination if given the opportunity, which may have important implications for continued VZV vaccination. Although no interclade vaccine-wild-type recombinant strains were found, intraclade recombinants were frequently detected in clade 2, which harbors the vaccine strains, suggesting that the vaccine strains have already been involved in recombination events, either in vivo or in vitro during passages in cell culture. Finally, previous partial and complete genomic studies have described strains that do not cluster phylogenetically to any of the five established clades. The additional VZV strains sequenced here, in combination with those previously published, have enabled us to formally define a novel sixth VZV clade. IMPORTANCE Although genetic recombination has been demonstrated to frequently occur for other human alphaherpesviruses, herpes simplex viruses 1 and 2, only a few ancient and isolated recent recombination events have hitherto been demonstrated for VZV. In the present study, we demonstrate that VZV also frequently undergoes genetic recombination, including strains belonging to the clade containing the vOKA strain.

Varicella-zoster virus infections of the central nervous system – Prognosis, diagnostics and treatment

Both varicella and herpes zoster that are caused by varicella-zoster virus (VZV), are associated with central nervous system disease. Since the introduction of polymerase chain reaction, the opportunity to detect the virus in cerebrospinal fluid (CSF) has improved dramatically. As a result VZV is diagnosed as one of the most common viruses causing CNS disease and it has become evident that this disease includes a wide spectrum of different CNS manifestations. The most evaluated CNS manifestations are encephalitis which is associated with both varicella and herpes zoster and, cerebellitis which occurs predominantly in children with varicella. Other manifestations have been less widely investigated. The incidence of cerebrovascular disease caused by VZV has been only scarcely studied and, in addition, some data indicate that vasculitis might also be involved in other VZV CNS manifestations such as herpes zoster-associated encephalitis. For this reason, VZV CNS infection must be suspected in several CNS syndromes and diagnostics should be based on CSF analysis for detection of VZV DNA by PCR and/or intrathecal antibody production. The prognosis is reported as favourable in children but few follow-up studies are available. Moreover, in adults, the prognosis is reported to be good in overall terms, but later studies indicate more serious neurological sequelae including cognition. Despite considerable mortality and morbidity, so far also in vaccinating countries, few treatment studies are available. Further treatment studies including assessments of neurological and cognitive sequelae, are warranted.

Varicella-Zoster Virus (VZV) Glycoprotein E Is a Serological Antigen for Detection of Intrathecal Antibodies to VZV in Central Nervous System Infections, without Cross-Reaction to Herpes Simplex Virus 1

ABSTRACT Herpes simplex virus 1 (HSV-1) and varicella-zoster virus (VZV) cause serious central nervous system (CNS) diseases that are diagnosed with PCR using samples of cerebrospinal fluid (CSF) and, during later stages of such infections, with assays of intrathecal IgG antibody production. However, serological diagnoses have been hampered by cross-reactions between HSV-1 and VZV IgG antibodies and are commonly reported in patients with herpes simplex encephalitis (HSE). In this study we have evaluated VZV glycoprotein E (gE) as a new antigen for serological diagnosis of VZV-induced CNS infections. Paired samples of CSF and serum from 29 patients with clinical diagnosis of VZV CNS infection (n = 15) or HSE (n = 14), all confirmed by PCR, were analyzed. VZV gE and whole VZV were compared as antigens in enzyme-linked immunosorbent assays (ELISAs) for serological assays in which the CSF/serum sample pairs were diluted to identical IgG concentrations. With the gE antigen, none of the HSE patients showed intrathecal IgG antibodies against VZV, compared to those shown by 11/14 patients using whole-VZV antigen (P < 0.001). In the patients with VZV infections, significantly higher CSF/serum optical density (OD) ratios were found in the VZV patients using the VZV gE antigen compared to those found using the whole-VZV antigen (P = 0.001). These results show that gE is a sensitive antigen for serological diagnosis of VZV infections in the CNS and that this antigen was devoid of cross-reactivity to HSV-1 IgG in patients with HSE. We therefore propose that VZV gE can be used for serological discrimination of CNS infections caused by VZV and HSV-1.

Recombinant glycoprotein E produced in mammalian cells in large-scale as an antigen for varicella-zoster-virus serology.

A recombinant glycoprotein E (gE) from varicella-zoster virus (VZV) was generated and produced in Chinese Hamster Ovary (CHO) cells, in the development of a specific antigen for analysis of IgG antibodies to VZV. Several stable gE-secreting clones were established and one clone was adapted to growth in serum-free suspension culture. When the cells were cultured in a perfusion bioreactor, gE was secreted into the medium, from where it could be easily purified. The recombinant gE was then evaluated as a serological antigen in ELISA. When compared to a conventional whole virus antigen, the VZV gE showed similar results in ELISA-based seroprevalence studies of 854 samples derived from blood donors, students, ischemic stroke patients and their controls, including samples with border-line results in previous analyses. Eight samples (0.9%) were discordant, all being IgG-negative by the VZV gE ELISA and positive by the whole virus ELISA. The sensitivity and specificity of the VZV gE ELISA were 99.9% and 100%, respectively, compared to 100% and 88.9% for the VZV whole virus ELISA. The elderly subjects showed similar reactivities to both antigens, while VZV gE gave lower signals in the younger cohorts, suggesting that antibodies to gE may increase with age. It was concluded that the recombinant VZV gE from CHO cells was suitable as a serological antigen for the detection of IgG antibodies specific for VZV.

Imported Case of Lassa Fever in Sweden With Encephalopathy and Sensorineural Hearing Deficit

We describe an imported case of Lassa fever with both encephalopathy and bilateral sensorineural hearing deficit. Absence of fever during hospitalization, initially nonspecific symptoms, and onset of hearing deficit in a late stage of disease probably contributed to delayed diagnosis (14 days after admittance to hospital). The pathogenesis of neurological manifestations of Lassa fever is poorly understood and no specific treatment was given. A total of 118 personnel had close contact with the patient, but no secondary cases occurred. This case highlights the importance of considering Lassa fever as a differential diagnosis in patients with recent travel to endemic areas.

Cerebrospinal fluid viral load and biomarkers of neuronal and glial cells in Ramsay Hunt syndrome.

Reactivation of varicella zoster virus (VZV) can manifest with facial palsy diagnosed as Ramsay Hunt Syndrome (RHS) or Ramsay Hunt Syndrome zoster sine herpete (RHS‐ZSH). These syndromes are associated with poor prognosis despite treatment with antivirals and corticosteroids. Concentrations of biomarkers such as neurofilament protein (NFL), S‐100β protein and glial fibrillary acidic protein (GFAp) have previously been measured in cerebrospinal fluid (CSF) to assess neuronal damage and glial pathology. We employed immunochemical methods to measure concentrations of NFL, S‐100β protein and GFAp in CSF from patients with RHS (n = 15) and RHS‐ZSH (n = 13) diagnosed by detection of VZV DNA in the CSF by quantitative PCR, and compared with a control group (n = 52). The biomarker concentrations were correlated with CSF viral load and outcome measured by House‐Brackmann score. NFL and GFAp concentrations were increased compared with controls (P = 0.008 and P = 0.04), while S‐100β levels were decreased. This pattern was more pronounced in patients with RHS compared to the patients with RHS‐ZSH (NS and P = 0.028). The amount of viral DNA in CSF correlated with increased GFAp (P = 0.003) and NFL (P = 0.006). No correlations were found between biomarker concentrations and patient outcome. Patients with facial palsy caused by VZV had biochemical signs of neuronal damage and astrogliosis. High amounts of viral DNA may be associated with the degree of damage on neuronal and astroglial cells. Prospective studies are warranted to elucidate the association of elevated biomarkers in the CSF and outcome assessed by more sensitive tests.

Absence of Nosocomial Transmission of Imported Lassa Fever during Use of Standard Barrier Nursing Methods

Nosocomial transmission of Lassa virus (LASV) is reported to be low when care for the index patient includes proper barrier nursing methods. We investigated whether asymptomatic LASV infection occurred in healthcare workers who used standard barrier nursing methods during the first 15 days of caring for a patient with Lassa fever in Sweden. Of 76 persons who were defined as having been potentially exposed to LASV, 53 provided blood samples for detection of LASV IgG. These persons also responded to a detailed questionnaire to evaluate exposure to different body fluids from the index patient. LASV-specific IgG was not detected in any of the 53 persons. Five of 53 persons had not been using proper barrier nursing methods. Our results strengthen the argument for a low risk of secondary transmission of LASV in humans when standard barrier nursing methods are used and the patient has only mild symptoms.