Ward KN on HHV-6 : clinical ramifications of chromosomal integration into telomeric regions in humans

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1: J Med Virol. 2008 Sep 23;80(11):1952-1958. [Epub ahead of print]

Human herpesvirus 6 integrates within telomeric regions as evidenced

by five different chromosomal Sites.

Nacheva EP, Ward KN, Brazma D, Virgili A, J, Leong HN, DA.

Department of Haematology, Royal Free & University College Medical

School (Hampstead Campus), London, United Kingdom.

Fluorescent in situ hybridization (FISH) was used to investigate the

chromosomal integration sites of human herpesvirus 6 (HHV-6) in

phytohemagglutinin-stimulated leukocytes and B lymphocytes from

Epstein-Barr virus transformed lymphoblastoid cell lines (LCLs). Five

different chromosomal integration sites were found in nine individuals.

Only one site was identified in each individual, each site was in the

vicinity of the telomeric region and was on either the p or q arm of

only one of the two chromosome homologues. The sites were 9q34.3,

10q26.3, 11p15.5, 17p13.3, and 19q 13.4, of which three have not been

previously identified. For 9q34.3 the site of integration was further

mapped using a locus-specific probe for 9q34.3 together with a

pan-telomeric probe and both co-localized with the HHV-6 signal.

Similarly an arm-specific telomeric probe for 19q co-localized with the

HHV-6 signal. It was therefore concluded that the site of integration is

actually within the telomere. The number of viral DNA copies/cell was

calculated in blood, LCL cells and hair follicles and was one or more in

every case for each of the nine individuals. This result was confirmed

by FISH where 100% of cells gave an HHV-6 signal. These findings add to

previous reports suggesting that integrated HHV-6 DNA is found in every

cell in the body and transmitted vertically. Finally, including our

data, worldwide seven different chromosomal sites of HHV-6 integration

have now been identified. Large epidemiological studies of chromosomal

integration are required to identify further telomeric sites,

geographical or racial variation and possible clinical consequences. J.

Med. Virol. 80:1952-1958, 2008. © 2008 Wiley-Liss, Inc.

PMID: 18814270

2: J Clin Microbiol. 2007 Apr;45(4):1298-304. Epub 2007 Jan 17.

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Human herpesvirus 6 DNA levels in cerebrospinal fluid due to primary

infection differ from those due to chromosomal viral integration and

have implications for diagnosis of encephalitis.

Ward KN, Leong HN, Thiruchelvam AD, Atkinson CE, DA.

Centre for Virology, Division of Infection and Immunity, Royal Free

and University College Medical School (UCL campus), Windeyer Institute

of Medical Sciences, 46 Cleveland Street, London W1T 4JF, United

Kingdom. k.n.ward@...

The prevalence and concentration of human herpesvirus 6 (HHV-6) DNA

in the cerebrospinal fluid (CSF) of the immunocompetent in primary

infection was compared with that in viral chromosomal integration.

Samples from 510 individuals with suspected encephalitis, 200 young

children and 310 older children and/or adults, and 12 other patients

were tested. HHV-6 DNA concentration (log(10) copies/ml) was measured in

CSF, serum, and whole blood using PCR. Serum HHV-6 immunoglobulin G

antibody was measured by indirect immunofluorescence. Primary infection

was defined by antibody seroconversion and/or a low concentration of

HHV-6 DNA (<3.0 log(10) copies/ml) in a seronegative serum. Chromosomal

integration was defined by a high concentration of viral DNA in serum

(>/=3.5 log(10) copies/ml) or whole blood (>/=6.0 log(10) copies/ml).

The prevalences of CSF HHV-6 DNA in primary infection and chromosomal

integration were 2.5% and 2.0%, respectively, in the young children (<2

years) and 0% and 1.3%, respectively, in the older children and/or

adults. The mean concentration of CSF HHV-6 DNA in 9 children with

primary infection (2.4 log(10) copies/ml) was significantly lower than

that of 21 patients with viral chromosomal integration (4.0 log(10)

copies/ml). Only HHV-6B DNA was found in primary infection, whereas in

viral integration, 4 patients had HHV-6A and 17 patients HHV-6B. Apart

from primary infection, chromosomal integration is the most likely cause

of HHV-6 DNA in the CSF of the immunocompetent. Our results show that

any diagnosis of HHV-6 encephalitis or other type of active central

nervous system infection should not be made without first excluding

chromosomal HHV-6 integration by measuring DNA load in CSF, serum,

and/or whole blood.

Publication Types:

* Research Support, Non-U.S. Gov't

PMID: 17229866 [PubMed - indexed for MEDLINE]

PMCID: PMC1865851

3: Curr Opin Infect Dis. 1998 Aug;11(4):425-30.

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Infections due to the herpesvirus group in immunocompromised patients.

Ward KN.

Department of Infectious Diseases, Imperial College School of

Medicine, Hammersmith Hospital, Du Cane Road, London W12 0NN, UK.

Herpesvirus infections remain a significant cause of morbidity and

mortality in iatrogenically immunosuppressed individuals despite the

considerable progress achieved in recent years by the use of anti-viral

drugs. Human cytomegalovirus is particularly important in this context

and major advances have been made towards understanding the basis of

cytomegalovirus latency and persistence. At the same time efforts

continue to develop optimal virus detection in immunosuppressed

individuals with a view to improving current therapeutic policies, and

to define the emerging problem of anti-viral drug resistance. Finally,

the evidence is gathering that the newest additions to the human

herpesvirus family, i.e. human herpesviruses-6, -7 and -8, are

significant pathogens in the immunosuppressed.

PMID: 17033405 [PubMed]

4: J Med Virol. 2007 Jan;79(1):45-51.

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The prevalence of chromosomally integrated human herpesvirus 6

genomes in the blood of UK blood donors.

Leong HN, Tuke PW, Tedder RS, Khanom AB, Eglin RP, Atkinson CE, Ward

KN, Griffiths PD, DA.

Division of Infection and Immunity, Centre for Virology, Hampstead

Campus, Royal Free and University College Medical School, London, UK.

A lesser-recognized form of human herpesvirus 6 (HHV-6) persistence

is integration of the viral genome in a host chromosome and high viral

copy numbers in blood or sera are characteristic of this phenomenon. A

cross-sectional study was performed to determine the frequency of high

HHV-6 viral loads in whole blood (>6 log(10) copies/ml) in a population

of blood donors in London, UK. Blood samples from 500 anonymized blood

donors were collected from one donation center, DNA extracted, and

quantitative realtime PCR used to measure viral load. Four samples

(0.8%) were found to have high viral copy numbers of HHV-6 (median 6.7

log(10) copies/ml; range 6.5- 6.9 log(10) copies/ml). Cellular DNA was

also quantitated using qRT-PCR for beta-globin. By comparing these two

results, we calculated that there were between two and five copies of

HHV-6 present per cell in these four donors. The median viral load

detected in plasma from the four individuals was 3.8 log(10) copies/ml

(range 3.5-4.0 log(10) copies/ml). All samples were HHV-6 variant B. In

addition, a retrospective analysis of all diagnostic blood samples

performed for HHV-6 in our center showed a prevalence of 2.9% of high

viral loads characteristic of integration. In conclusion, high viral

copy numbers of HHV-6, representing a population of viral integration,

is detected in 0.8% of UK blood donors. The presence of high HHV-6 viral

loads in healthy normal individuals reiterates the need to consider the

confounding effect of HHV-6 viral integration in any laboratory

diagnosis of HHV-6 infection.

PMID: 17133548 [PubMed - indexed for MEDLINE]

5: J Med Virol. 2005 Aug;76(4):563-70.

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Erratum in:

* J Med Virol. 2005 Sep;77(1):144.

Unexpected occasional persistence of high levels of HHV-6 DNA in

sera: detection of variants A and B.

Ward KN, Thiruchelvam AD, Couto-Parada X.

Department of Virology, Royal Free and University College Medical

School, Windeyer Institute of Medical Sciences, 46 Cleveland Street,

London, United Kingdom. k.n.ward@...

Previously it was thought that in the immunocompetent human

herpesvirus-6 [HHV-6] DNA was present transiently in serum during early

primary infection but not thereafter. In this study, HHV-6 serum IgG

avidity was detected by immunofluorescence and HHV-6 variants A/B

[HHV-6A/B] serum DNA by semi-quantitative PCR [titre-log(10) copies/ml]

in: (a) young children <3 years old from an encephalitis Survey, and a

control Anonymised Serum Bank and ( children/adults referred for

diagnosis. The results showed that 11 out of 15 children [all <2 years]

with primary infection proven by seroconversion had transient low levels

of serum HHV-6B DNA [mean titre 2.6]. However, 3.3% (6/184) of Survey

Children had significantly higher levels [mean titre 5.3; 2 HHV-6A; 4

HHV-6B; P < 0.001]. Similarly high level serum DNA [mean titre 4.0; 4

HHV-6A; 6 HHV-6B] was found in 1.5% (10/653) of the Serum Bank Children.

Moreover, seven young children <3 years old [four Survey Children and

three referred for diagnosis] had high titre serum HHV-6 DNA [mean 4.8]

persisting i.e., in all available samples [median 186 days]. Three older

children >3 years old and 4 adults [3 of whom were the mothers of 3 of

the young children with persisting HHV-6] also had persisting high titre

viral DNA [mean 4.2; median 108 days]. Thus in contrast to acute primary

infection, where only HHV-6B DNA is found transiently, both HHV-6A and B

DNA persist in serum at high titre in occasional individuals of all

ages. The significance of this newly described phenomenon in relation to

diagnosis, clinical consequences and congenital infection are discussed.

© 2005 Wiley-Liss, Inc.

Publication Types:

* Research Support, Non-U.S. Gov't

PMID: 15977239 [PubMed - indexed for MEDLINE]

6: Arch Dis Child. 2005 Jun;90(6):619-23.

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Human herpesviruses-6 and -7 each cause significant neurological

morbidity in Britain and Ireland.

Ward KN, s NJ, Verity CM, E, Ross EM.

Department of Virology, Royal Free and University College Medical

School, Windeyer Institute of Medical Sciences, London, UK.

k.n.ward@...

BACKGROUND: Primary human herpesvirus-6 and -7 (HHV-6/-7) infections

cause febrile illness sometimes complicated by convulsions and rarely

encephalopathy. AIMS: To explore the extent of such HHV-6 and -7 induced

disease in young children. METHODS: In a three year prospective study in

Britain and Ireland, 205 children (2-35 months old) hospitalised with

suspected encephalitis and/or severe illness with fever and convulsions

were reported via the British Paediatric Surveillance Unit network.

Blood samples were tested for primary HHV-6 and -7 infections. RESULTS:

26/156 (17%) of children aged 2-23 months had primary infection (11

HHV-6; 13 HHV-7; two with both viruses) coinciding with the acute

illness; this was much higher than the about three cases expected by

chance. All 26 were pyrexial; 25 had convulsions (18 status

epilepticus), 11 requiring ventilation. Median hospital stay was 7.5

days. For HHV-6 primary infection the median age was 53 weeks (range

42-94) and the distribution differed from that of uninfected children;

for HHV-7, the median was 60 weeks (range 17-102) and the distribution

did not differ for the uninfected. Fewer (5/15) children with primary

HHV-7 infection had previously been infected with HHV-6 than expected.

CONCLUSIONS: Primary HHV-6 and HHV-7 infections accounted for a

significant proportion of cases in those <2 years old of severe illness

with fever and convulsions requiring hospital admission; each virus

contributed equally. Predisposing factors are age for HHV-6 and no

previous infection with HHV-6 for HHV-7. Children with such neurological

disease should be investigated for primary HHV-6/-7 infections,

especially in rare cases coinciding by chance with immunisation to

exclude misdiagnosis as vaccine reactions.

Publication Types:

* Research Support, Non-U.S. Gov't

PMID: 15908629 [PubMed - indexed for MEDLINE]

PMCID: PMC1720457

7: J Clin Virol. 2005 Mar;32(3):183-93.

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The natural history and laboratory diagnosis of human

herpesviruses-6 and -7 infections in the immunocompetent.

Ward KN.

Centre for Virology, Department of Infection, Royal Free and

University College Medical School (UCL campus), Windeyer Institute of

Medical Sciences, 46 Cleveland Street, London W1T 4JF, UK.

k.n.ward@...

BACKGROUND: Human herpesviruses-6 and -7 (HHV-6/7) are widespread in

all populations. In some individuals HHV-6 is found integrated into

human chromosomes, which results in a high viral load in blood. HHV-6

variant B (HHV-6B) and HHV-7 primary infections, although usually

silent, not infrequently cause the childhood exanthem roseola infantum

and are sometimes accompanied by neurological illness. HHV-6 variant A

(HHV-6A) is not associated with any disease. OBJECTIVES: The present

review focuses on the immunocompetent individual and considers the

epidemiology of the two viruses and their role as human pathogens. It

discusses the importance of satisfactory diagnostic tests to distinguish

them, compares those currently available, and recommends how best to

differentiate primary from persistent infection in each case. RESULTS:

It is explained that at the present time antibody avidity

immunofluorescence tests are the most reliable discriminators of the two

types of infection. In primary infection these tests can be supplemented

by PCR for viral DNA in blood but careful interpretation is required for

HHV-6 in view of the high persistent viral DNA load seen with

chromosomal integration. Since the contribution of primary HHV-6 and -7

infections to the burden of severe neurological illness in young

children is only now emerging as significant, the need to test for these

viruses in such cases is stressed. CONCLUSIONS: 1. Primary HHV-6/7

infections must be distinguished from persistent infections. 2.

Chromosomal integration of HHV-6 requires urgent study. 3. HHV-6A/B must

be distinguished in clinical situations. 4. Where serious neurological

disease/encephalitis is temporally related to immunisation it is

particularly important to test for HHV-6/7 primary infection since

otherwise the condition might wrongly be diagnosed as a vaccine

reaction. 5. Because less is currently known about HHV-7 and HHV-6A than

HHV-6B, future studies should concentrate on the former two. 6.

Improvements in diagnostic tests are required for each virus.

Publication Types:

* Review

PMID: 15722023