Opportunistic Infections in Immunocompromised Patients
Moderator: Dr. Paul Hofman
Section 3 -
Diagnostic Virology of EBV and Adenovirus in Transplant Recipients
Randall T. Hayden
Immunocompromised patients are subject to a variety of infections which may produce marked differences
in epidemiology, morbidity and mortality, compared to immunocompetent individuals. Disease spectrum and
severity may depend heavily on a patient's underlying medical condition, the cause and degree of
immunosupression. Viral infection represents a primary concern in such populations, with particularly
increased frequency and consequences of infection among those with diminished cell-mediated immunity.
Thus, patients with primary or secondary immune deficiencies may be at great risk for symptomatic and
often severe infection with a wide array of viral pathogens. While the spectrum of viral infections in
this group is broad, many laboratory diagnostic challenges are shared among them. Most such issues can
be well-illustrated with only a few examples. Epstein-Barr virus (EBV) and adenovirus (ADV) will be
discussed in this context.
- 90-95% adult seropositivity rate worldwide
- Spread by oropharyngeal secretions
- Infects B lymphocytes
- Associated with several forms of neoplasia, including
post-transplant lymphoproliferative disorder (PTLD)
- Associated with allogeneic transplantation and
- Mortality approaching 90% in some series
- Risk Factors
- Non-enveloped DNA viruses
- 51 serotypes, six subgroups
- Variable homology within and between subgroups
- Increasing association with disease, mortality in
immunocompromised individuals, particularly BMT patients
- hemorrhagic cystitis, enteritis, hepatitits,
- Prevalence of ADV 5-37%
- Invasive disease in 30-80%
- Risk factors
Laboratory Diagnostics – Challenges
- Speed of detection
- Clinical Correlation
While the identification of adenovirus in clinically infected tissues can be accomplished by
culture-based methodologies, as well as by direct morphologic evidence of the infectious agent (i.e.,
viral inclusions), the identification of Epstein-Barr Virus in such specimens is largely based on
histopathologic characterization of typical lymphoproliferative processes, in conjunction with techniques
such as immunohistochemistry or in situ hybridization.
In the case of adenovirus, findings include typical histopathologic evidence of infection, most often
described in hepatic or pulmonary lesions. These lesions variably include features such as necrosis,
epithelial hyperplasia, and inflammatory infiltrates. The latter may of course vary depending on the
degree of immunosuppression in a given case. Intra-nuclear inclusions are usually present and are
typically amphophilic or basophilic and smudgy in appearance. Diagnosis may be confirmed through the use
of either immunohistochemistry or in situ hybridization. Adenovirus may
also be isolated from various specimens using conventional cell culture methods, with the production of
characteristic cytopathic effects and confirmation of diagnosis by immunoflourescent methods. These
techniques are specific, but have limited sensitivity and may require prolonged incubation of culture for
definitive results. Antigen detection methods are more rapid, but may also exhibit limited sensitivity.
Epstein Barr Virus, as noted above, may provoke a lymphoproliferative process, hyperplasic or
neoplastic in nature, without direct morphologic evidence of viral proliferation. The virus may be
specifically detected in characteristic lesions using either immunohistochemistry or in situ hybridization. Immunohistochemistry targets include Epstein Barr Virus
nuclear antigen (EBNA), latent membrane protein 1 (LMP-1), or Epstein-Barr-encoded RNAs (EBERs). In situ hybridization can also target one of several sequences, including EBNA,
the BamH1 W repetitive sequence, or EBERs.
Viral isolation from EBV-related lesions is not often performed, requiring co-cultivation of the tissue
of interest together with umbilical cord lymphocytes. This procedure is of limited practicality in the
clinical laboratory and its sensitivity may also be suboptimal.
Detection of Systemic Infections
While the detection of EBV and Adenovirus can be accomplished in the setting of clinically or
histopathologically evident end-organ infection, this may represent a relatively late stage of disease.
Systemic detection of these viruses, through routine surveillance techniques, may allow preemptive action
at an earlier stage, with a greater potential for preventing serious or life-threatening illness. The
tools available for such testing include all of the usual laboratory methodologies. However,
traditional, phenotypic techniques, such as viral culture, are often too insensitive to detect the
concentrations of virus circulating in the early phases of active infection. Nucleic acid amplification
methods, such as PCR, hold great promise for the early detection of infection, as well as for the
assessment of treatment efficacy and the determination of therapeutic endpoints. The remainder of the
discussion will deal with the use of molecular methodologies including their strengths and limitations,
and the particular advantages of real-time target amplification methods. Also discussed will be assay
selection, design, and verification of performance, as well as ongoing assay performance and the use of
test results in projecting clinical disease.
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