Diagnosis of Viral Hepatitis

Hepatic inflammation, acute or chronic, may be drug-induced, toxin-related, or a manifestation of viral infection of the liver.

Aetiology

Acute hepatitis

• • Acute viral hepatitis
    • Hepatitis A virus (HAV)
    • Hepatitis B virus (HBV) ± Delta
    • HCV, HDV (delta agent), HEV
  • Hepatitis with systemic [viral] infection
    • Cytomegalovirus (CMV)
    • Epstein Barr Virus (EBV)
    • Herpes simplex
    • Enteroviruses: Coxsackie virus; Echo virus
    • Yellow Fever
    • Q Fever
    • Non-viral: Toxoplasmosis; Leptospirosis
  • Alcoholic hepatitis — heavy alcohol consumption over an extended period of time (i.e. subacute)
    • other chemical hepatitis
      • carbon tetrachloride (CCl4) — dry-cleaning solvent
      • mushroom toxins — approximately 100 of more than 5000 species are poisonous to humans
  • drug-induced liver disease
    • Paracetamol — dose-dependent
    • Isoniazid (INH) — complicates prolonged anti-tuberculous therapy
    • Halothane — free radicals formed upon hepatic metabolism and also subsequent sensitisation [ now replaced by newer halogenated anesthetics]
    • Chlorpromazine (and other phenothiazines)
    • Idiosyncratic – e.g. Phenytoin: rare acute idiosyncratic severe or even fatal
    • Tetracycline

Chronic hepatitis

• chronic viral hepatitis
• autoimmune chronic active hepatitis — often asymptomatic chronic transaminitis
  • ~ 50% positive for anti-nuclear antibody (ANA)
  • ± smooth muscle antibody (SMA) positive
  • anti-mitochondrial antibody positive (primary biliary cirrhosis)
• chronic persistent hepatitis
• chemicals, drugs
• Wilson’s disease

Conceptually, Autoimmune Chronic Active Hepatitis (AI-CAH) evolved out of work from the Italian Association for the Study of the Liver who, in September 1989, recognised a syndromic mimic of acute viral hepatitis in patients with otherwise symptomless jaundice and who, left untreated, went on to liver failure. But, to their delight, the group discovered that these patients responded remarkably promptly to low-dose prednisolone. In contradistinction to elevated IgM levels in patients with primary biliary cirrhosis (PBC), these patients showed an elevated IgG production. These patients’ liver biopsies, moreover, showed a characteristic morphological picture: piecemeal necrosis; portal tract plasma-cell infiltrate; with or without “liver cell rosettes” (indicative of sub-massive necrosis); and a frequently coexistent cirrhosis. Within this then still maturing field of immunology, autoantibody characterisation rendered to this subgroup the markers of anti-nuclear antibody (ANA), anti-smooth muscle (SMA) antibody, anti-liver-kidney microsomal (LKM) antibody, liver-specific membrane lipoprotein (LSP), and asialoglycoprotein receptor (ASGP-R) antibody; in contrast to the anti-mitochondrial antibodies (AMA) seen in over 95% of PBC patients or the gastric-parietal cell (GPC) antibodies seen, for instance, in Pernicious Anaemia but with variable elevation of thyroid microsomal autoantibodies and antibodies against either reticulin or thyroglobulin. [5]

Further characterisation confirmed another subgroup of patients, not only without overt viral markers (i.e. they are HBV Ab and HCV Ab negative) but none either of these typical serological markers mentioned above. To this group of patients, the term “cryptogenic CAH” has been attached. Far more commonly seen in women, by a ratio of 11:1, especially those of HLA B8 and DR3 allotypes, cryptogenic CAH otherwise shows a similar chronic transaminitis and liver histomorphology with AI-CAH and, further, responds to immunosppressives. Some suggest this syndrome characterises a “burnt-out” form of CAH.

“Most weight attaches to the autoantibody markers, which are predominantly to antigens of nuclei and smooth muscle. The multiple nuclear antigenic reactants include DNA (rarely), histones and lamins, and the smooth muscle reactant is the cytoskeletal filament, F-actin. Autoantibody to neutrophil cytoplasmic antigen (ANCA) is also seen. In a variant now called autoimmune hepatitis type 2, the marker antigen is enriched in liver and kidney microsomes (LKM). The LKM antigen has been identified as a particular cytochrome P450 isoform, CYP450 2D6. There is suspected to be a liver-specific autoantigen, perhaps membrane-associated, and one candidate is the asialoglycoprotein receptor (ASGP-R); however this to a degree lacks fine disease specificity for association with autoimmune hepatitis. Long-term treatment with prednisolone, combined with azathioprine, is highly effective”. [6]

“Hepatitic” enzymes

Transaminases: catalyse the transfer of an alpha-amino group to ketoglutaric acid to produce oxaloacetic and pyruvic acids. They are elevated in most liver diseases but highest in hepatitis, especially that caused by viruses and toxins; though values do not necessarily correlate with prognosis.

• aspartate aminotransferase (AST / SGOT)
  • aspartate substrate
  • heart, muscle, kidney, brain and liver
  • mitochondrial and cytosolic
• alanine aminotransferase (ALT / SGPT)
  • alanine substrate
  • liver-specific enzyme
  • cytosolic

Consider, as a differentiating factor, the AST / ALT ratio which, while low in viral hepatitis, is usually > 2 in alcoholic hepatitis and rarely above 300 IU/L in absolute terms.

Once acute hepatitis is suspected, aim to identify the cause taking a thorough history and examination, including possible drug and toxin exposure, travel history, sore throat and adenopathy suggestive of EBV, history of ethanol ingestion, stigmata of chronic liver disease.

Many viruses can cause diffuse inflammation of the liver but the term viral hepatitis is generally reserved for that caused by a handful of diverse viruses with a relatively long incubation that show hepatotropy—a predilection for hepatocytes. These viruses are successively labelled as Hepatitis A through to Hepatitis E, while Epstein-Barr (EBV), Cytomegalovirus (CMV) and Yellow Fever viruses, for instance, are generally considered among the non-hepatitic viruses that nevertheless can cause a hepatitis.

Hepatitis A Virus

Hepatitis A is a single-stranded RNA picornavirus that causes acute, and occasionally fulminant, food- and water-borne hepatitis but no chronic disease. It is known for epidemics in children and young adults of underdeveloped countries where more than 75% of adults have been exposed, conferring to them life-long immunity.

“Being a small non-enveloped virus, it [Hepatitis A] can withstand environmental stress and maintain its infectivity. The incubation period is usually 30 days, but can be between 15 to 50 days”. [4]

Because shedding begins before, and ends a few days after, symptoms develop, most patients are non-infectious at presentation. While usually self-limiting and acute, prolonged viral replication can, however, occur in those with pre-existing liver disease or in the immunosuppressed. [4]

In acute infection, request HAV IgM, and although not routinely performed post-vaccination, total HAV will detect immunity to hepatitis A virus.

Hepatitis B virus

The outer surface coat of Hepatitis B hepadnavirus replicates excessively in the cytoplasm while its core of circular double-stranded DNA (with DNA polymerase) replicates within hepatocyte nuclei and is transmitted parenterally from blood rather than through casual exposure through infected bodily fluids, such as saliva and semen, while also often passed vertically from mother to foetus. The long incubation period varies from 45 to 180 days.

Interestingly, Hepatitis B infection is often associated with extrahepatic (autoimmune) disease—such as polyarteritis nodosa (PAN) and other connective tissue diseases (CTDs), membranous GMN, and essential mixed cryoglobulinemia.

“The majority of adults will recover with clearance of the virus from the blood after a few months. The development of chronic hepatitis B occurs when there is a failure of the immune response to eradicate the virus”. [4]

Such a failure occurs in about 5-10% of patients, and viral persistence in these patients is a marker of either chronic hepatitis (CAH) or the inactive carrier state, both providing for a worldwide reservoir of infection. This endemicity varies from region to region: from less than 0.5% in North America and northern Europe; to more than 10% in some regions of the Far East. Hepatitis B infection is associated with a propensity for cirrhosis and hepatocellular carcinoma, usually as late sequelae of infection. Initial testing of HepBsAg is invoked in antenatal screening, among other groups. At risk groups should be offered vaccination.

Hepatitis C virus

There are six major subtypes of the Hepatitis C, a single-stranded RNA flavivirus, varying in geography and virulence and showing mutability of amino acid sequence over time (quasi-species). Transmission invariably involves contaminated blood, with sexual and vertical transmission occurring far less commonly.

“The incubation period can range from two weeks to six months, though a six to nine week incubation period is common”. [4]

Because of its propensity (70%) to chronicity, Hepatitis C is frequently associated with, and exacerbates, alcoholic liver disease but otherwise remains asymptomatic yet indolently progressing (often presenting decades later) to a cirrhosis in 20-30% of patients, a cirrhosis not infrequently associated with hepatocellular carcinoma. HCV antibodies persist for life, regardless of viral eradication, cure, or persistence. [4]

Infection chronicity behooves upon medical practitioners to test and treat for Hepatitis C with direct-acting antivirals that achieve up to 95% cure rates.

“Persons at risk of contracting a blood borne disease should be tested for hepatitis C as should persons with chronic liver disease or abnormal liver enzymes”. [4]

Screen these people for hepatitis C antibody (HCV Ab). If positive,
request a molecular (HCV PCR) test and genotyping. Consider molecular testing also in patients presenting acutely, perhaps toward the end of the window incubation period, in whom serology may still be negative. [4]

Offer Hepatitis A and Hepatitis B vaccination in patients with, or at-risk of, Hepatitis C. There is no vaccine against Hepatitis C virus.

Hepatitis D virus

Hepatitis D is actually a defective RNA “delta antigen” that replicates only in presence of HBV, usually as a super-infection of chronic Hepatitis B, in which case hepatocytes will contain delta particles coated with HBsAg. Its prevalence varies widely with pockets of endemicity, particularly in parenteral drug users, but far less prevalent in the homosexual community, for instance, than HBV. It is not commonly seen in Australia. [4]

Hepatitis E virus

An enterically transmitted RNA virus, hepatitis E infection is often linked to faecal contamination of water supply in south America, Eurasia, and northern Africa, conferring an epidemiology not unlike HAV epidemics.

“Although classically thought to be a disease seen in travellers returning from the developing world, it has been increasingly described in Europe especially in relation to pig farming and may be underdiagnosed in Australia”. [4]

While there is no chronic disease or carrier state associated with hepatitis E infection, it is a serious disease in pregnant women and may lead to fulminant liver failure in up to 20% of them. [4] Diagnosis is made by antibody testing where other serology remains negative, and confirmed by HEV PCR.

Pattern of clinical presentation

Incubation period: asymptomatic.

Patients may remain asymptomatic but otherwise constitutional symptoms precede jaundice by 1-2 weeks.

1. Prodrome (Pre-icteric) Phase: non-specific symptoms for 3-10 days
   • fever / headache / malaise / lassitude
   • profound anorexia; nausea and vomiting; diarrhoea
   • other GI symptoms / Abdominal pain (RUQ)
   • ± Hep B Prodrome: arthritis (arthralgia), rash (urticaria), angioedema
2. Icteric Phase: patient starts to feel better despite
   1. darkening urine / pale stools / jaundice which peaks within 1-2 weeks
   2. tender hepatomegaly (soft, smooth edge) ± cholestasis
   3. mild splenomegaly (10-20%)
   4. ± enlarged cervical nodes
3. Convalescent Phase: 2-12 weeks
   1. 2-4 weeks where appetite returns and jaundice fades to a spontaneous resolution some 4-8 weeks after symptom onset (longer in Hepatitis B)
   2. hepatomegaly and abnormal LFTs often persist for some time
   3. occasional recrudescence

Anicteric hepatitis, presenting as flu-like illness without the jaundice, is seen especially with Hepatitis C and in children with HAV infection. (Note: ascites, encephalopathy etc. are not  features of uncomplicated viral hepatitis.)

Diagnosis of Acute Viral Hepatitis

Use the whole spectrum of clinical assessment at your disposal: i.e. history, examination, biochemistry (LFTs), haematology (Prothrombin Time), imaging, and serology.

1. Clinical
   1. characteristic symptomatology
   2. contact history
   3. drugs / alcohol history
      1. IVDU, blood transfusion, STI
2. Serum Biochemistry (LFTs)
   1. bilirubin
   2. serum transaminases (ALT & AST) x 10 Normal (often > 1000 IU)
      1. ALT (≥ 400 IU/L) > AST >> ALP (only moderately elevated), usually with hyperbilirubinemia; but levels correlate poorly with clinical severity
      2. asymptomatic chronic transaminitis in many patients with hepatitis
         • up to 72% histology c/w chronic active hepatitis (CAH)
         • ~ 50% positive anti-nuclear antibody (ANA) ± smooth muscle antibody (SMA)
           • exclude HBV / HCV but consider also drugs, Wilson’s disease, sclerosing cholangitis, alpha-antitrypsin deficiency, non-alcoholic steatohepatitis (NASH)
   3. albumin – normal
   4. FBC – normal / low WCC
3. Serological tests: diagnosis in 99% cases
   1. HBsAg, anti-HBc; IgM – anti-HAV
      1. Above negative in 30% → anti-HCV* (selected patients): anti-HepC Ab can take 3-4 months to seroconvert: specificity ~ 50% in acute, higher in chronic
         1. If negative → anti-HDV (selected patients)
4. Imaging
   1. ultrasonography if suspect biliary obstruction
   2. biliary scan (e.g. DISHIDA) if suspect acute cholecystitis
5. Prothrombin Time
   1. test of “liver cell function” if suspect severe disease; good guide to prognosis
   2. suggests fulminant hepatitis
6. Liver Biopsy
   1. only when atypical, prolonged (> 6 months) course or diagnosis uncertain
   2. Carriers HBcAg / HBV DNA positive

Acute Viral Hepatitis – Differential Diagnosis

1. Hepatitis / hepatic necrosis
   1. Alcoholic hepatitis
   2. Drug hepatitis
   3. Systemic infections: CMV, EBV, Q Fever, Septicaemia, Pneumonia
   4. Ischaemic hepatitis
   5. Chronic hepatitis
   6. Wilson’s disease
2. Jaundice
   1. Unconjugated hyperbilirubinaemia
      1. Gilbert’s syndrome (3%  of population)
      2. haemolysis
   2. Conjugated hyperbilirubinaemia
      1. cholestasis – gallstones, pancreatic or bile duct carcinoma, cholestasis of pregnancy
3. Abdominal Pain
   1. especially acute cholecystitis

Hepatitis serology

In the acute setting, the following markers should be tested:

• anti-HAV IgM
• HBsAg
• anti-HBc IgM
• anti-HCV

If any of these are positive, further testing will differentiate acute from past or chronic infection, accordingly:

Hepatitis A (HAV)

Test for antibody to the HAV:

• HAV (only present acutely and no clinically available serological test)
• Anti-HAV IgM: acute infection – develops early, peaking at 1-2 weeks after jaundice and diminishes within several weeks, followed by;
• Anti-HAV IgG: past infection – confers protective, usually life-long, immunity

Hepatitis B (HBV)

HBV avails us of surface, core, and e antigen-antibody systems for testing.

Surface antigen-antibody:

Surface antigen characteristically appears during the incubation period and implies infectivity,  and disappears during convalescence.

• HBsAg: acute / chronic infection –
  • occasionally transient –> False Negative testing during window period

The corresponding protective antibody appears week or months later, after clinical recovery, and usually persists for life.

• Anti-HBs (HBsAb):corresponding protective antibody
  • past infection (recovered)
  • the sole serologic marker after HBV vaccination

In 5-10% of patients, however, HBsAg persists and the corresponding antibodies do not develop. These patients become asymptomatic carriers of the virus or develop chronic hepatitis.

Core antigen-antibody:

HBcAg reflects the viral core and is most easily detectable within liver cells than blood. The corresponding antibody generally appears at onset of clinical illness and gradually diminishes over years to decades. Its presence, in combination with anti-HBs, indicates recovery from infection. It is also seen in those 5-10% of patients (see above) who do not mount a protective surface antibody response — i.e. those who go on to become carriers or develop chronic hepatitis. In acute infection, anti-HBc is mainly IgM which acts as sensitive (and occasionally the only) marker of acute hepatitis B infection during a window between the disappearance of the surface antigen (HBsAg) and the appearance of its corresponding surface antibody (anti-HBs).

• Anti-HBc IgM: acute infection

In chronic infection, the IgG form of anti-HBc predominates.

• Anti-HBc IgG: past or chronic infection

e antigen=antibody:

HBeAg is a protein derived from the viral core present only in HBsAg-positive serum and indicates active viral replication and greater infectivity.

• HBeAg: acute / chronic infection (prognostic / management implications)

In contrast, presence of its corresponding antibody suggests lower infectivity.

• Anti-HBe: past or chronic infection (prognostic / management implications)

e-antigen markers are useful then as prognostic indicators, with chronic disease more common in those with e-antigen and less common in those with the corresponding anti-HBe.

Hepatitis B virus DNA is not routinely tested for.

• HBV DNA (replicative phase): acute / chronic infection

Note: antibodies to Hepatitis D [Delta] Ab (anti-HDV) levels should be measured if serologic tests confirm HBV and infection is severe. Anti-HDV implies active infection but may not be detectable until weeks after the acute illness.

Hepatitis C (HCV)

Serum antibody to HCV (anti-HCV) is not protective and invariably implies active infection, usually appearing within about two weeks of acute infection.

• Anti-HCV Ab: acute / chronic / past (recovered or cured) infection

Sometimes the presence of this antibody in serum is delayed, in which case HCV-RNA will still be positive. Only in a small proportion of patients does anti-HCV reflect merely prior exposure with clearance of virus, rather than active infection, in which case ALT and AST levels usually remain normal. If in doubt, measure HCV-RNA.

• HCV-RNA: acute / chronic infection

Hepatitis E (HEV)

The test for anti-HEV IgM is not routinely available but, in the presence of endemic exposure and compatible clinical findings is suggestive for Hepatitis E virus infection.

“Where it is unclear what tests should be performed, request “Hepatitis Serology” and the laboratory will assign testing based on the clinical notes. All of the serological testing can be performed on serum samples. PCR testing requires a dedicated tube”. [4]

Other IgM assays to consider in hepatitis:

• Cytomegalovirus (CMV)
• Epstein-Barr (EBV)
• Herpes simplex (HSV)
• Varicella-zoster (VZV)

And others still, such as those for Measles virus, Rubella virus, and Coxsackie viruses.

Brief Guide to management Acute Viral hepatitis

1. Symptomatic / Supportive
   1. bed rest
   2. diet, alcohol avoidance (avoid fats)
   3. treatment of pruritus etc.
2. Prevention of cross-infection
   1. Recall HAV: faecal-oral route
   2. HBV and HCV: blood infective (no faecal/oral spread)
   3. sexual transmission – especially HBV: safe sex or abstinence
   4. gamma globulin – HBIG (Hep B immune globulin)
      1. within 24 hours (up to 1 week) of exposure
         1. 80-90% prevention of disease
      2. can be given together with vaccine
      3. give to neonate on day of delivery of Hep B positive mother
   5. Hep B vaccination
3. Interferon

Natural History of Acute Viral hepatitis

Chronic active hepatitis is treated with antivirals, interferon, and even liver transplantation.

Complications of Viral hepatitis

1. Benign
   1. relapse – similar to initial infection
   2. prolonged cholestasis – especially after Hepatitis A (good outlook)
   3. persistent hepatitis (chronic)
   4. unconjugated hyperbilirubinaemia
   5. post-hepatitis syndrome
2. Serious
   1.  fulminant hepatitis (massive hepatic necrosis)
   2. subacute hepatitis (bridging hepatic necrosis ± submassive hepatic necrosis)
   3. chronic active hepatitis (chronic aggressive hepatitis)
   4. cirrhosis
   5. hepatocellular carcinoma (HBV and HCV)

When in doubt, ultrasound-guided liver biopsy will confirm the diagnosis. Histopathology is remarkably similar regardless of the specific viral cause: patchy cell loss; acidophilic necrosis; mononuclear infiltrate; and evidence of regeneration but with preservation of the reticulin (supportive) architecture. Ground-glass hepatocytes, chock-full of HBsAg, usually represent chronic HBV infection. Complete recovery of histological architecture occurs unless necrosis crosses entire acini, but even still most patients with this so-called bridging necrosis recover fully.

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Further Reading

• Natural history of hepatitis B virus infection
• Hepatitis B and Primary Care Providers – ashm
• All You Wanted to Know About Hepatitis B
• Decision-Making in HCV – ashm
• HCV Treatments Quick Reference Tool – ashm
• Epidemiology and treatment of autoimmune hepatitis. Francque S, Vonghia L, Ramon A, Michielsen P. Epidemiology and treatment of autoimmune hepatitis. Hepat Med. 2012;4:1–10. Published 2012 Mar 16. doi:10.2147/HMER.S16321

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References

1. Kumar, Sonal. “Overview of Acute Viral Hepatitis”. Merck, Sharpe and Dohme (MSD) Manual, Professional Edition. Last full review/revision Oct 2019 | Content last modified Oct 2019. Available at https://www.msdmanuals.com/en-au/professional/hepatic-and-biliary-disorders/hepatitis/overview-of-acute-viral-hepatitis?query=hepatitis.
2. Servoss, Julie C et al. “Diagnostic Approach to Viral Hepatitis.” Viral Hepatitis, Blackwell Publishing Ltd, 2007, pp. 50–64, doi:10.1002/9780470987131.ch4.
3. Pendle, Stella. “The A-B-Cs of Diagnosing Viral
   Hepatitis in General Practice”. Pathology Focus 2, July 2018: 5-7. https://www.clinicallabs.com.au/doctor/gp-services/newsletters/pathology-focus-july-2018/.
4. Mackay, I. R. (1990) Auto‐immune (lupoid) hepatitis: An entity in the spectrum of chronic active liver disease*. Journal of Gastroenterology and Hepatology. [Online] 5 (3), 352–359.
5. Features of specific types of CAH Autoimmune hepatitis – Immune Response (2018). Available at: https://www.arthritisresearch.us/immune-response/features-of-specific-types-of-cah-autoimmune-hepatitis.html (Accessed: 16 April 2020).