1. Case 5: Not as Ezee as ABC Educational Workshops 2015
We are grateful to Emanuela Pelosi, Consultant Medical Virologist,
University Hospitals Southampton NHS Foundation Trust
and Manoj Valappil, Honorary Consultant Virologist, Newcastle for
composing these cases.

2. Case 5 a

3. Patient’s history 56 year old male End Stage Liver Disease
Decompensated cirrhosis caused by non-alcoholic steatohepatitis
MELD score 26 (19.6% 3 month mortality)
: Orthotopic liver transplant
CMV IgG status: Recipient + / Donor _
Immune suppressive treatment instituted to prevent graft rejection
Prednisolone 15mg od (weaned and stopped in Jan 14)
Azathioprine
Tacrolimus (later substituted with sirolimus)
MELD: Model for End Stage Liver Disease
scoring system for assessing severity of liver disease. Useful in determining prognosis and prioritizing for receipt of liver transplant. MELD uses the patient's values for serum bilirubin, creatinine and INR to predict survival.
In interpreting the MELD Score, 3 month mortality is:
40 or more 71.3% mortality
30– % mortality
20– % mortality
10– % mortality
< % mortality
Child Pugh Score Grade C

4. Oct 2013 – July 2014
Patient well with normal liver and renal functions
Management of CMV and EBV infections
No evidence that CMV prophylaxis has been instituted
CMV and EBV DNA are monitored once/month in order to implement pre-emptive treatment in case of reactivation
CMV DNA remains undetectable
EBV VL fluctuates between undetectable and very low levels (< 500 IU/ml)

5. August 2014
Transient rise in ALT levels over a period of 2 weeks with peak of 142 U/L
Thought to be related to systemic reaction to
Lidocaine (local anesthetic used to excise a small squamous cell carcinoma on a forearm) or to
ACE inhibitors (treatment for hypertension is modified)
However, in view of transplant need to exclude rejection
Liver biopsy: no evidence of graft rejection
CMV DNA remains undetectable as well as EBV DNA

6. DIFFERENTIAL DIAGNOSIS?
Develops mild jaundice and acute transaminitis
ALT value is very raised (989 U/L)
He is very well
DIFFERENTIAL DIAGNOSIS?

7. 1.12.14 Serum sample tested for viral hepatitis markers
Hep A IgM negative
HBsAg negative
HCV IgG negative
CMV DNA undetectable
EBV DNA undetectable
Hep E IgM highly positive
IgG low positive
Viral Load 9 million IU/ml
Diagnosis of HEV infection
ALT value 795 U/L

8. Diagnosis of HEV infection requires confirmation by testing a second sample
Hep E infection confirmed on a new sample
HEV IgG and IgM detected
HEV RNA detected at 12 million IU/ml
HEV genotyped in the reference laboratory:
Genotype 3, closely related to HEV strains circulating in the UK
ALT value 349 U/L
Questions
Source of infection?
What to do next?

9. Answers To question 1 (Source of infection?)
Most likely food related infection, as patient had not received blood products in the 2 months preceding onset
To question 2 (what to do next?)
HEV VL monitoring in blood to see whether the infection is spontaneously cleared over time

10. End Feb. 2015
Immune suppressive treatment is decreased (tacrolimus stopped and sirolimus reduced) due to altered kidney function.
Two important results are generated
Significant rise in ALT level
Spontaneous drop in HEV VL (NB result from ref lab available only 7 days later)
As the patient still HEV RNA positive 3 months after onset, the case fulfills the definition of chronic HEV infection

11. Questions
3. How do you interpret these changes? 3a. rise in ALT level 3b. drop in HEV VL 4. Should the liver transplant team wait and see whether the patient is able to clear the infection? 5. What is the best antiviral option in case they decide to treat chronic hepatitis E?

12. Answers
To question 3 (How do you interpret drop in HEV VL and rise in ALT level and)
Reduction of immune suppression has triggered
antiviral immune response and
acute graft rejection (histological evidence of acute graft rejection in liver biopsy)
At this point immune suppressive regimen is intensified (high-dose
methylprednisolone) to prevent rejection

13. Answers
To question 4 (Should the liver transplant team wait and see whether the patient is able to spontaneously clear the infection?)
The transplant team had initially decided to monitor HEV VL and LFTs hoping for spontaneous clearance of infection
Spontaneous clearance was more likely to occur when immune suppression was reduced due to renal toxicity
Indeed, decrease of immunosuppression may be considered as 1st line treatment option, in which case patients must be closely monitored (risk of graft rejection)
The concomitant onset of acute graft rejection reverses the decision and immune suppression is intensified (high-dose m-prednisolone is added)
At this point spontaneous clearance of HEV infection is unlikely to occur and the transplant team decides to treat HEV infection with an antiviral drug

14. Answer
To question 5 (What is the best antiviral drug option to treat chronic hepatitis E?)
Our patient is started on oral Ribavirin 600mg bd
Background information about anti-HEV treatment options
Peg IFN-alpha & Ribavirin  only drugs currently available
Ribavirin is used as monotherapy or in combination with IFN
Limited scientific evidence for the use of these drugs
case series & in vitro studies
not validated in controlled trials yet
Use of IFN in the transplant setting may increase the risk of graft rejection
Reference: Antiviral therapy in chronic hepatitis E: a systematic review.
J Viral Hepatitis 2015 Mar 11. doi: /jvh.12403

15. Answer
To question 5 (What is the best antiviral drug option to treat chronic hepatitis E? ) (cont)
Mechanism by which ribavirin inhibits HEV replication
Largely unknown
Depletion of cellular GTP pools: predominant mechanism in vitro
However unclear to what extent hepatic GTP levels are decreased in vivo
Other proposed mechanisms include
Immune-modulatory effect
Modulation of IFN-stimulated gene expression
Lethal mutagenesis forcing the virus into an error catastrophe,
Interference with viral methyltransferase activity,
Direct inhibition of the viral polymerase
Inhibition of eukaryotic initiation factor 4E interfering with cap-dependent translation
Reference: Debing et al, Antimicrobial Agents and Chemotherapy January 2014 Volume 58 Number 1

16. Answer
To question 5 (What is the best antiviral drug option to treat chronic hepatitis E?) (cont)
Reasons why our transplant team decided to use ribavirin monotherapy
Ribavirin is well tolerated and effective in clearing HEV infection
Main side effect  anaemia (during treatment some patients may require recombinant erythropoietin or, less frequently, blood transfusion)
This treatment option also recommended by Public Health England (Virus Reference Unit)
Effectiveness of ribavirin monotherapy in solid organ transplant recipients with prolonged viremia has been recently reviewed:
Ribavirin for chronic hepatitis E virus infection in transplant recipients. N Engl J Med 2014 Mar 20;370(12): (retrospective, multicenter case series).

17. Question 6
How would you assess if ribavirin treatment is successful in treating hepatitis E virus infection?

18. Ribavirin stopped 30.6.15 (treated for 3 months)
Answer to question 6 - HEV monitoring in blood and stools - Two negative HEV PCR in stool, once HEV RNA has become undetectable in blood, indicate clearance of infection
Ribavirin stopped (treated for 3 months)
Prednisolone stopped
Patients remains HEV RNA negative (August 2015)
HEV IgG and IgM are still detectable at high titres (August 2015)
CMV and EBV VLs remain undetectable (CMV) or at low-level (EBV)

19. Prednisolone and Ribavirin started

20. Summary of case 5a
Investigation of a patient with acute transaminitis due to HEV infection post liver transplant
Acute Hepatitis E progressed to chronic persistent HEV infection due to immunosuppression
Source of HEV infection likely related to food, the most common source of infection in the UK and in other developed countries (genotype 3)
Reduction in immunosuppressive regimen due to tacrolimus renal toxicity triggered antiviral immune response and graft rejection reactions; the latter event prompted the decision to intensify immune suppression and to start ribavirin monotherapy.
Clearance of HEV infection was confirmed by testing stool samples by HEV PCR.

21. Case 5 b

22. History November 2011 23 year old female
ALL, chemotherapy, in remission
Philadelphia positive, high risk for remission
March 2012
Full intensity Allogeneic HSCT
Unrelated donor

23. April 2012 Increased ALT, AST and Bilirubin
HBsAg, HCV RNA & HAV RNA: Not detected
CMV, EBV, Adenovirus and HHV-6 DNA: Not detected
?GvHD
?other pathology

25. Progress Cyclosporine related TTP Changed to Tacrolimus
No significant improvement
HEV test added

26. May 2012 HEV IgM Reactive, HEV IgG negative HEV RNA: Detected
Q1) Is this acute HEV?
Q2) What is the source?

27. Answers Q1) Is this Acute HEV? Q2) What is the source?
Possible acute HEV
Need to differentiate from chronic persistent replication that can occur in immunocompromised patients
Test previous samples stored in the lab
Q2) What is the source?
Incubation period ~ 9 weeks
Common sources in the UK: under cooked pork products
Transfusion transmitted HEV described
Was hospitalised for most of this period
Unlikely from hospital catering
Decision to investigate

28. Progress Q1) Stored recipient samples Feb-March (per-transplant):
HEV IgM negative, HEV RNA detected.
HEV RNA also detected in a sample stored for an ALL trial
Interpretation: Not acute HEV, consistent with Chronic HEV. Asymptomatic due to immune suppression, immune mediated liver damage after engraftment post BMT
Q2) Donor: HEV IgM, IgG, HEV RNA: not detected
Not transmitted from bone marrow donor
Had transfusions Nov 2011 – Feb 2012
?Transfusion Transmitted Infection (TTI) – notify NHSBT

29. Management No guidelines Consensus practice:
Reduction of immunosuppression
Follow up for up to 3 months to see if spontaneous clearance
If no clearance – Ribavirin
Plan:
Reduce immunosuppression
Follow up HEV viral load and serology

30. Follow up HEV genotype 3 (mostly indigenous/UK infection)
Other genotypes – mostly travel associated
HEV RNA detected at high level (10^7 copies/ml)
No IgG sero-conversion

31. NHSBT look back Had multiple transfusions from multiple donors
2 donors had HEV viraemia at the time of donation.
Both donors had HEV genotype 3
Phylogenetic analysis:
HEV strain detected in the patient was closely related to the one from donor 1
Transfusion Dec 2011

32. Genotype 3 Donor 2 Donor 1 Recipient Genotype 2 Genotype 1 Genotype 4
27.3 25 20 15 10 5
Nucleotide Substitution per 100 residues

34. Interpretation: HEV transmission via blood transfusion from donor 1

35. June 2013 Fully engrafted HEV viral load still high, HEV IgG negative
Stopped all immunosuppression
Follow up:
HEV IgG seroconversion
Fall in HEV viral load
HEV RNA Not detected (July 2013)

37. Another case
Another Haematology patient (non-BMT) presented with acute HEV
Multiple transfusions in the past
Had transfusion from the same donor!

38. Summary of case 5b
Investigation of a patient with abnormal LFTs post Bone marrow transplant
Identified as chronic persistent HEV due to immunosuppression
Source of HEV infection traced back to a blood donor who was viraemic at the time of donation
With engraftment and stopping of immunosuppression patient mounted adequate immune response to clear HEV without Ribavirin treatment
Another patient who received blood product from the same donor developed acute HEV

39. Learning points Sources of HEV infection in the UK
Blood products: this source of infection cannot be prevented as blood products not currently screened for HEV
Food: preventable source  avoidance of undercooked game meat and pork products
HEV diagnostic tests
In our two cases HEV infection has been identified by antibody testing
However, it is worth underlining that negative HEV IgG and IgM results do not exclude HEV infection in immunocompromised patients, particularly in those who are profoundly immunosuppressed
Therefore HEV PCR should be considered in cases of unexplained acute transaminitis or persistent deranged ALT values in immunocompromised patients with negative HEV antibody results

40. Learning points (cont.)
3. Outcome of HEV infections
In immunocompetent adults, acute HEV infection is a self-limiting illness
In immunocompromised patients HEV can establish chronic persistent infection associated in some cases with rapid progression to cirrhosis within 1-2 years of infection
Reduction of/stopping immunosuppression or improved immune function (e.g. HSCT patients with good graft function) may help spontaneous clearance of HEV infection
In patients with ongoing immunosuppression (e.g. immune suppression cannot be stopped due to risk of graft rejection), HEV is less likely to clear spontaneously
Spontaneous clearance of HEV relies on T-cell mediated immune response
HEV infections have been also linked to a variety of extrahepatic manifestations, such as CNS disease both in immunocompetent and in immunocompromised patients . There was no evidence of such manifestations in our two patients

41. Learning points (cont.)
4. Treatment
Immunosuppressed patients may be followed up to monitor spontaneous clearance (unless otherwise indicated)
If antiviral treatment is considered, Ribavirin monotherapy currently represents the best treatment option
It is well tolerated and it is effective at a relative low dose
Treatment failure may occur

42. Learning points (cont.)
5. Monitoring of HEV VL during ribavirin treatment (no guidelines, current practice):
Ribavirin treatment is associated with clearance of HEV in
chronically infected immunosuppressed patients
During Ribavirin treatment:
Monitor HEV viral load in blood until negative
Testing stools once HEV RNA has become undetectable in blood (anecdotal evidence of treatment failure if treatment stopped whilst still excreting in stool)
Stop Ribavirin after 2 negative HEV RNA results in stool