Bone Marrow Transplantation, (1998) 22, 587–589
 1998 Stockton Press All rights reserved 0268–3369/98 $12.00
Case report
Acyclovir-resistant herpes simplex virus infections in a bone marrow
transplant population
JM Darville1,2, BE Ley3, APCH Roome2 and ABM Foot3
1
Department of Pathology and Microbiology, University of Bristol, Bristol; 2Public Health Laboratory, Bristol; and 3Department of
Paediatric Oncology, Bristol Royal Hospital for Sick Children, Bristol, UK
Summary:
Over a 3-month period, four patients who had received
unrelated donor (UD) bone marrow transplants (BMT)
presented with severe mucocutaneous herpes simplex
virus (HSV) infection while receiving acyclovir (ACV)
prophylaxis. Sensitivity testing of the isolates revealed
three to be acyclovir-resistant and in one patient the
infection was also characterised by a marked failure to
respond to foscarnet (phosphonoformic acid). The
emergence of ACV-resistant HSV infections in them-
selves is a new and challenging problem, and yet a far
greater problem will become evident if these infections
develop resistance to non thymidine kinase dependent
therapy.
Keywords: acyclovir-resistant herpes simplex virus
infections; bone marrow transplantation
Herpes simplex virus (HSV) reactivates in 70–80% of sero-
positive recipients of allogeneic transplants, causing haem-
orrhagic mucocutaneous lesions which can add substan-
tially to the morbidity of BMT. These infections are
preventable with prophylactic acyclovir (ACV).1,2 The
mode of action of acyclovir includes phosphorylation by
viral thymidine kinase (TK) and then its incorporation into
the viral DNA, thus terminating extension of the DNA
chain. Resistance is usually due to loss of TK expression
(TK−) and a correlation exists between the frequency with
which resistant viruses are isolated and the degree of
immunosuppression in the host.3,4 ACV-resistant HSV
infections causing clinical problems in BMT patients have
previously been documented,5 as have resistant herpes virus
infections generally.6
We present a review of four patients who had received
unrelated donor bone marrow transplants for the treatment
of haematological malignancies and presented over a 3-
month period between July and September 1995. They each
had serological evidence of previous HSV infection at the
time of transplant (complement fixing titres ranging from
Correspondence: Dr JM Darville, Public Health Laboratory, Myrtle Road,
Kingsdown, Bristol, BS2 8EL, UK
Received 28 November 1997; accepted 10 May 1998
http://www.stockton-press.co.uk/bmt
24–64) and all four developed recurrent HSV lesions while
receiving ACV prophylaxis. The standard regimen for ACV
prophylaxis in UD BMT at the time was commenced at
day −4 and continued until 6 months post-transplant, but
was prolonged in the case of active graft-versus-host dis-
ease (GVHD). In patients at risk of cytomegalovirus
(CMV) reactivation, the ACV was commenced at
500 mg/m2 intravenously (i.v.) three times per day until day
30 and then changed to an oral dosage (200 mg five times
per day). In those patients not deemed at risk for CMV,
ACV was given orally throughout. The implications of the
emergence of resistance in this population of immunosup-
pressed patients will be further discussed.
Case report
Patient A, an 8-year-old girl with biphenotypic leukaemia
developed mouth ulcers, culture positive for HSV-1, 5 days
post-transplant (isolate A, day +5). She was changed from
her oral ACV prophylaxis to high-dose intravenous ACV
(10 mg/kg three times daily) for 2 weeks, to which she
responded and was then converted back to oral therapy
(400 mg × 5/day). However, on developing diarrhoea a
week later the mouth ulcers recurred and on this occasion
they were successfully treated with a course of foscarnet.
The ulcers were again culture positive for HSV-1 but the
later isolate was not tested for ACV susceptibility.
Patient B, a 13-year-old female with T cell ALL,
developed HSV mouth ulceration from 6 days post-trans-
plant while she was already receiving high-dose i.v. ACV
(500 mg/m2 three times daily) and she was therefore
changed to treatment with foscarnet from day +17 to which
she responded (isolate B, day +19). After 3 weeks she was
converted back to oral ACV (400 mg × 5/day) without
further reactivation.
Patient C, a 25-year-old female with AML, first
developed HSV-positive mouth ulcers concurrent with the
development of GVHD 16 days post-transplant whilst
receiving low-dose oral ACV prophylaxis. These ulcers
responded to conversion to 5 days of high-dose i.v. ACV
therapy (500 mg/m2 tds). Eight months post-transplant,
whilst still on steroid therapy and low-dose oral ACV
prophylaxis, she had a further recurrence of HSV mouth
ulcers associated with a flare-up of GVHD. She was treated
 ACV-resistant HSV in BMT patients
JM Darville et al
588
with high-dose i.v. ACV for 3 weeks, but without response
on this occasion (isolate C, day +206). During this admis-
sion she developed haemolytic uraemic syndrome with
cerebral involvement and died shortly after.
Patient D, a 28-year-old male with AML, had prolonged
immunosuppression post-transplant with severe GVHD. Six
months post-transplant, while continuing on low-dose
prophylactic oral ACV, he developed oral HSV lesions
which progressed across his face despite high-dose i.v.
ACV therapy (isolate D, day +206). On conversion to fos-
carnet, the facial lesions regressed but did not clear. Due
to severe toxicity the foscarnet was stopped after 3 weeks,
with concomitant flare-up and progression of the facial
lesions through further treatment with ACV and famciclo-
vir. A second course of foscarnet was attempted, on this
occasion without clinical response, and was terminated
after 10 days because of unacceptable side-effects. Despite
ongoing therapy with i.v. ACV and penciclovir he still had
spreading HSV-1 culture-positive erosive lesions across his
face and left middle finger. Foscarnet was recommenced
on day 340 but after 3 weeks there was no clinical response.
At the time of his death from bacterial sepsis 12 months
post-transplant, he still had clinical HSV disease and at
post-mortem there was evidence of both local and dissemi-
nated HSV infection.
Laboratory data
Samples from the four patients were inoculated into tubes
of vero and human embryo fibroblast cells. On the develop-
ment of HSV-specific cytopathic effect (CPE) the isolates
were typed by direct immunofluorescence using mono-
clonal antibodies (Syva Microtrak, Milton Keynes, UK): all
were found to be HSV-1. Duplicate cultures of each were
allowed to progress to complete CPE and the titre of virus
in the supernatant of each was measured by plaque assay
on vero cells under a methylcellulose overlay. The sensi-
tivity of each strain was determined by a plaque reduction
method, incorporating dilutions of ACV (Wellcome, Lon-
don, UK) into the overlay. SC16, a standard strain7 of HSV-
1 was used as the sensitive control and R16 (a TK− deriva-
tive of SC16) as the resistant control. Sensitivities were
expressed as the ACV concentration which reduced plaque
number by 50% (EP50). The sensitivities (Figure 1) of the
100
90
80
% Plaques remaining
70
60
50
40
30 SC16 R16 A ing ACV treatment of recurrent infections in immuno-
20 B C D compromised patients may add to the number of neurones
10
0
0.1 0.33 1 3.3
Acyclovir µ g/ml
Figure 1 Sensitivity of HSV isolates to acyclovir.
controls were ⬍0.1 ␮g/ml (SC16) and 1–3.3 ␮g/ml (R16).
Those of the clinical isolates were ⬍0.1 ␮g/ml (isolate A),
1–3.3 ␮g/ml (isolates B and C) and ⬎10 ␮g/ml (isolate D).
Discussion
These four patients all developed significant HSV infec-
tions whilst receiving ACV post-UD BMT and in three
cases the isolate tested resistant in vitro. Isolate A proved
to be as sensitive in vitro as the sensitive control and the
clinical failure of the ACV treatment in this case was prob-
ably due to poor absorption of the oral preparation second-
ary to mucositis and diarrhoea. Isolates B and C showed
patterns of resistance similar to that of the resistant control,
while isolate D was highly resistant. These results relate
closely to the clinical course of the HSV infections in
these patients.
ACV resistance usually occurs through the loss of
expression of TK but mutants producing less TK, altered
TK and altered DNA polymerase have also been reported.
The TK− strains have little or no virulence in the immuno-
competent, but TK− ACV resistance in clinically apparent
HSV infections is more common in all immunocompro-
mised patients.8 However, the failure of clinical response
to foscarnet, as seen in patient D, has previously been
described only in HIV-infected individuals: in 26 patients
with AIDS an 81% clinical response rate was seen in ACV-
resistant HSV infection.9
The difference in degree of resistance found in isolate D
in contrast to isolates B and C may reflect different mech-
anisms of resistance, while the failure of treatment with
foscarnet suggests that it is a DNA polymerase mutant. Fos-
carnet is structurally unrelated to ACV and acts by
inhibiting DNA polymerase by binding to its pyrophosph-
ate site. Cross-resistance does not occur with HSV strains
having TK-based resistance, but could with DNA poly-
merase-based resistance involving the pyrophosphate site.
Since immunocompromised patients are susceptible to
infection with TK− resistant strains, there is a possibility
that in these patients TK− mutants could establish or re-
establish latency, with adverse consequences for the treat-
ment of future recurrent disease. It is also possible that
transmission to other immunocompromised patients could
occur. In mouse models it has been reported that ACV
resistant TK− mutants of HSV can produce low levels of
TK as a result of a frameshift mechanism and that these
are able to reactivate from latency in the mouse trigeminal
ganglion.10 Furthermore, it has also been reported that TK−
HSV strains can reactivate in AIDS patients.11 ACV treat-
ment has been shown to reduce both the numbers of neu-
rones infected and the viral copy number per cell in ganglia
after primary HSV infection.12 TK− mutants generated dur-
latently infected, so effective alternative treatment of this
group of patients may be important in respect of future
10 disease episodes.
The importance of the interaction of immune response
with ACV therapy in the recovery from HSV infections as
reported by Wade et al4 is also seen in the cases reported

here, although in that study ACV was used intermittently.
Patient B developed resistant herpes infection early in the
post-transplant period while patients C and D both had
GVHD.
The commercial availability of other TK-dependent
agents such as famciclovir has not solved the problem of
ACV resistance and the development of resistance to ther-
apies not TK-dependent is an emergent problem in the
transplant population. It is clearly important to have an
effective battery of anti-HSV chemotherapeutic agents both
to treat current episodes and also to prevent problems with
future recurrent disease. The agent (S)-1-[3-hydroxy-2
(phosphonylmethoxy)-propyl] cytosine (HPMPC or
cidofovir), which has selective activity against a broad
range of DNA viruses including HSV, is an example of an
alternative drug which has been reported to be successful
in the topical treatment of ACV/foscarnet-resistant HSV
infection.13 It is not TK-dependent but acts directly on the
viral DNA polymerase, presumably by binding to other
than the pyrophosphate site. The intravenous preparation
has recently become licensed and may indeed prove bene-
ficial in such cases as described here, although there is
some concern that it may prove to be nephrotoxic if used
systemically.
Until the current biological sensitivity tests are replaced
by rapid molecular methods, decisions to alter antiviral
regimens will be based on clinical criteria, although it
should be borne in mind as in case A that clinical failure
does not always reflect virological resistance. The authors
suggest, therefore, the following clinical strategy. Patients
developing HSV lesions while on oral ACV should be
changed to high-dose i.v. ACV. If lesions persist after a
week or if the patient is already on i.v. ACV, then replace-
ment with foscarnet is advised. If after a further 10 days
there is no improvement a trial of cidofovir may be of
value. However, there is still an urgent need for more alter-
native, effective therapies to be developed for the small but
growing cohort of patients with ACV-resistant HSV
disease.
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