Cheryl S. Toi

Changes in patterns of hospitalized children with varicella and of associated varicella genotypes after introduction of varicella vaccine in Australia

Background: Varicella in children, although usually mild, can cause hospitalization and rarely death. This study examined patterns of hospitalized children with varicella, and associated varicella genotypes, in 4 tertiary children’s hospitals throughout Australia before and after varicella vaccine was introduced. Methods: We obtained coded data on discharge diagnoses from each hospital before (1999 to 2001) and after (2007 to 2010) varicella vaccine introduction in 2005, adding active surveillance to capture clinical features, complications and immunization history in the latter period. Varicella vesicles were swabbed, and genotyping of varicella strains was performed by real-time polymerase chain reaction amplification. Results: Overall, a 68% reduction in coded hospitalizations (varicella, 73.2% [P < 0.001]; zoster, 40% [P = 0.002]) occurred post-vaccine introduction. Of children with detailed clinical data (97 varicella and 18 zoster cases), 46 (40%) were immunocompromised. Only 6 of 32 (19%) age-eligible immunocompetent children were immunized. Complications, most commonly secondary skin infections (n = 25) and neurologic conditions (n = 14), occurred in 44% of children. There were no deaths; but 3 immunocompetent unimmunized children had severe multiple complications requiring intensive care. All strains genotyped were “wild-type” varicella, with Clade 1 (European origin) predominating. Conclusions: After the introduction of varicella vaccine, coverage of greater than 80% at 2 years of age was achieved, with varicella hospitalizations reduced by almost 70%. Of hospitalized children age-eligible for varicella vaccine, 80% were unimmunized, including all cases requiring intensive care.

Differentiation between vaccine and wild-type varicella-zoster virus genotypes by high-resolution melt analysis of single nucleotide polymorphisms

BACKGROUND The analysis of single nucleotide polymorphisms (SNPs) of varicella-zoster virus (VZV) has enabled differentiation between wild-type genotypes from the Oka vaccine strain (V-Oka). OBJECTIVES To genotype VZV strains in Australia using high-resolution melt (HRM) analysis of SNPs in five gene targets. STUDY DESIGN Extracted DNA from 78 samples obtained from patients with chickenpox and zoster were genotyped by HRM analysis of SNPs in five open reading frames (ORFs): 1 (685 G>A), 21 (33725 C>T), 37 (66288 G>A), 60 (101464 C>A) and 62 (106262 T>C) using a double-stranded (ds) DNA saturating dye, LC Green Plus. RESULTS For each genotype, melt curve temperature (Tm) shifts differentiated the nucleotide present at that locus (P<0.0001) with melting curve shifts between alleles ranging from 0.56 degrees C (ORF 37) to 3.34 degrees C (ORF 62). The most common genotypes detected were the European Type C (59%) and B (18%) strains. This was followed by the African/Asian Type A (14%) and Japanese J1 (9%), strains, both prevalent in the Northern Territory and Western Australia. CONCLUSIONS HRM analysis of SNPs showed that the European B and C genotypes were most prevalent in Australia, with genotypes A and J strains also present. HRM analysis using a dsDNA dye provides a useful tool in classifying varicella-zoster viruses.

Converting Mosquito Surveillance to Arbovirus Surveillance with Honey-Baited Nucleic Acid Preservation Cards.

Spatially and temporally accurate information about infectious mosquito distribution allows for pre-emptive public health interventions that can reduce the burden of mosquito-borne infections on human populations. However, the labile nature of arboviruses, the low prevalence of infection in mosquitoes, the expensive labor costs for mosquito identification and sorting, and the specialized equipment required for arbovirus testing can obstruct arbovirus surveillance efforts. The recently developed techniques of testing mosquito expectorate using honey-baited nucleic acid preservation cards or sugar bait stations allows a sensitive method of testing for infectious, rather than infected, mosquito vectors. Here we report the results from the first large-scale incorporation of honey-baited cards into an existing mosquito surveillance program. During 4 months of the peak virus season (January-April, 2014) for a total of 577 trap nights, we set CO2-baited encephalitis vector survey (EVS) light traps at 88 locations in South Australia. The collection container for the EVS trap was modified to allow for the placement of a honey-baited nucleic acid preservation card (FTA™ card) inside. After collection, mosquitoes were maintained in a humid environment and allowed access to the cards for 1 week. Cards were then analyzed for common endemic Australian arboviruses using a nested RT-PCR. Eighteen virus detections, including 11 Ross River virus, four Barmah Forest virus, and three Stratford virus (not previously reported from South Australia) were obtained. Our findings suggest that adding FTA cards to an existing mosquito surveillance program is a rapid and efficient way of detecting infectious mosquitoes with high spatial resolution.

Prevalence of varicella-zoster virus genotypes in Australia characterized by high-resolution melt analysis and ORF22 gene analyses.

DNA sequence variation analysis has divided varicella-zoster virus (VZV; Human herpesvirus 3) into distinct geographical clades: European, Asian, African and Japanese. These genotypes are becoming increasingly prevalent within regions atypical to their original source and there has been the suggestion of recombination between genotypes. Seventy-eight clinical isolates from hospitalized patients with varicella were collected in New South Wales, the Northern Territory, Western Australia and Victoria from 2006 to 2009. The wild-type strains and the vaccine strain (vOka) were differentiated by single nucleotide polymorphism detection using high-resolution melt analysis of five target genes (ORF1, -21, -37, -60 and -62), and by DNA sequence analysis of a 484 bp region of ORF22. Phylogenetic analysis showed that 46 % (36/78) of the clinical isolates were European clade 1 (C/E1) strains, 21 % (16/78) were European clade 3 (B/E2) strains, 12 % (9/78) were Asian/African clade 5 (A/M1) strains, 10 % (8/78) were clade 4 (J2/M2), 6 % (5/78) were clade 2 (J/J) and 5 % (4/78) belonged to the novel clade VI. No significant association was shown between VZV genotype and region, age or gender. Although European strains were most common, the results suggest an increase in African/Asian, Japanese and clade VI genotypes circulating in Australia.

Epstein-Barr Virus Genotypes and Strains in Central Nervous System Demyelinating Disease and Epstein-Barr Virus-Related Illnesses in Australia

Objectives: To identify Epstein-Barr virus (EBV) genotypes and strains in samples from individuals with and without a first diagnosis of central nervous system (CNS) demyelinating disease (a possible precursor to multiple sclerosis) and patients with EBV-associated diseases in Australia. Methods: Samples from 55 EBV DNA and serology positive subjects including individuals with (n = 17) and without (n = 21) a first clinical diagnosis of CNS demyelination and patients with EBV-related diseases (n = 17) were examined. EBV genotype and strain were identified by sequence mutations within the Epstein-Barr nuclear antigen-2 region (EBNA-2) using DNA sequence analysis. Results: Both EBV genotypes, A and B, were detected (genotype A, 54/55, 98.2%; genotype B, 1/55, 1.8%). Within genotype A, GD1 was the most commonly detected strain (42/54, 77.7%), followed by B95-8 (9/54, 16.7%) and M-ABA (3/54, 5.6%). Genotype B, strain AG876, was found in one individual with CNS demyelinating disease. Conclusions: EBV genotype A and the GD1 strain were the common EBV genotypes isolated from individuals with and without CNS demyelinating disease, and in subjects with various EBV-related diseases. Although disease-specific genotypes or strains were not identified, this study provides useful insights into the molecular epidemiology of EBV infection in Australia.

Varicella zoster virus quantitation in blood from symptomatic and asymptomatic individuals

Primary infection with varicella zoster virus (VZV) occurs in immunocompromised and immunocompetent individuals. Clinical and asymptomatic reactivation with shedding of infectious virus and viremia may occur. The prevalence of VZV viremia is unknown. The aim of this study was to detect VZV viremia and quantify VZV DNA using quantitative polymerase chain reaction (qPCR) in blood from different populations. A qPCR‐based method using EvaGreen® was used to quantify VZV DNA in 491 samples, including whole blood, plasma and buffy‐coat, from patients hospitalized with varicella‐associated disease (Group 1, n = 10) and three groups with no VZV disease: individuals with a first clinical diagnosis of central nervous system demyelination (Group 2, n = 213) with their age and sex‐matched controls (Group 3, n = 218); and HIV‐infected individuals (Group 4, n = 50). VZV‐specific IgG antibody titres were measured in Group 3. The proportion positive for viremia and mean detectable VZV DNA load (copies/ml) were: Group 1: 100% (10/10) and 4.6 × 106 ± 1.4 × 107; Group 2: 4% (9/213) and 1.5 × 103 ± 1.8 × 104; Group 3: 8% (17/218) and 1.1 × 103 ± 7.8 × 103; Group 4: 12% (6/50) and 7.7 × 101 ± 2.8 × 102. VZV DNA load and IgG titres were not significantly correlated (Group 3 only). VZV load in Group 1 was significantly elevated compared to Groups 2–4 (P < 0.001); the latter were not significantly different from each other (P = 0.05). VZV genotypes from clades 1–5 were identified in Group 1. VZV DNA was detected but at low frequency and viral load in both immunocompetent and immunocompromised individuals asymptomatic for VZV infection, compared to individuals with active VZV infection. J. Med. Virol. 85:1491–1497, 2013.

Seasonal activity, vector relationships and genetic analysis of mosquito-borne Stratford virus

There are many gaps to be filled in our understanding of mosquito-borne viruses, their relationships with vectors and reservoir hosts, and the environmental drivers of seasonal activity. Stratford virus (STRV) belongs to the genus Flavivirus and has been isolated from mosquitoes and infected humans in Australia but little is known of its vector and reservoir host associations. A total of 43 isolates of STRV from mosquitoes collected in New South Wales between 1995 and 2013 was examined to determine the genetic diversity between virus isolates and their relationship with mosquito species. The virus was isolated from six mosquito species; Aedes aculeatus, Aedes alternans, Aedes notoscriptus, Aedes procax, Aedes vigilax, and Anopheles annulipes. While there were distinct differences in temporal and spatial activity of STRV, with peaks of activity in 2006, 2010 and 2013, a sequence homology of 95.9%–98.4% was found between isolates and the 1961 STRV prototype with 96.2%–100% identified among isolates. Temporal differences but no apparent nucleotide divergence by mosquito species or geographic location was evident. The result suggests the virus is geographically widespread in NSW (albeit only from coastal regions) and increased local STRV activity is likely to be driven by reservoir host factors and local environmental conditions influencing vector abundance. While STRV may not currently be associated with major outbreaks of human disease, with the potential for urbanisation and climate change to increase mosquito-borne disease risks, and the possibility of genomic changes which could produce pathogenic strains, understanding the drivers of STRV activity may assist the development of strategic response to public health risks posed by zoonotic flaviviruses in Australia.

Severe and Complicated Varicella and Associated Genotypes 10 Years After Introduction of a One-Dose Varicella Vaccine Program

Background This national, sentinel prospective study aimed to identify children with severe hospitalized varicella, despite availability of universal 1-dose vaccination since 2005, and determine associations between virus genotypes and disease severity. Methods Children with varicella or zoster from 5 Paediatric Active Enhanced Disease Surveillance hospitals were enrolled. Lesions were swabbed for genotyping. Associations with disease severity were analyzed using multiple regression. Results From 2007 to 2015, 327 children with confirmed varicella (n = 238) or zoster (n = 89) were enrolled. Two hundred three (62%) were immunocompetent children; including 5 of 8 children who required intensive care unit management. Eighteen percent (36 of 203) of immunocompetent children had been previously vaccinated. Vaccinated children aged >18 months were less likely to have severe disease (9%; 5 of 56) than unvaccinated children (21%; 21 of 100; P = .05). Three of 126 children who had virus genotyping (2 immunocompromised) had varicella (n = 2) or zoster (n = 2) due to the Oka/vaccine strain. European origin clades predominated and were independently associated with more severe disease (odds ratio = 3.2; 95% confidence interval, 1.1- 9.5; P = .04). Conclusions Severe hospitalized varicella still occurs with a 1-dose varicella program, although predominantly in unvaccinated children. Most 1-dose vaccine recipients were protected against severe disease. Viral genotyping in complex hospitalized cases is important to assist in monitoring disease due to Oka-vaccine strain.

New Record of Wyeomyia mitchellii (Diptera: Culicidae) on Guam, United States

Abstract Wyeomyia (Wyeomyia) mitchellii (Theobald) (Diptera: Culicidae) was recovered for the first time on Guam, United States of America, in 2017. Larval specimens were collected from water-filled axils of bromeliads during a larval survey carried out in a residential neighborhood of the Chalan Pago/Ordot area. Native to the New World, Wy. mitchellii has likely made its way to the Pacific Islands through the possibly illegal import of ornamental bromeliads. While this mosquito does not represent a significant threat to public health, this finding highlights the vulnerability of the Pacific Islands to the introduction of exotic species, including mosquito species that may increase public health risks.