Chapter 14: Herpesviruses

VIROLOGY

All herpesviruses are morphologically similar, with an overall size of 180 to 200 nm. An example of a HSV virion is shown in Figure 14–1 as a representative virion structure for herpesviruses. The linear, double-stranded DNA genome and core proteins are encapsidated by an icosahedral capsid. The capsid is surrounded by the tegument, a relatively amorphous protein-filled region unique to herpesviruses. The tegument contains viral proteins and enzymes that play a structural role and many are required immediately for viral replication upon initial infection. Surrounding the tegument is a lipoprotein envelope originally derived from the nuclear membrane of the infected host cell. The envelope contains multiple viral glycoproteins that act as viral binding, fusion, and entry proteins.

Herpesvirus genomes range from 125 kbp (VZV) to 240 kbp (CMV) of DNA, and code from around 75 viral proteins to over 200. However, it is now clear from next-generation RNA sequencing and proteomics that the coding capacity is much more complex than originally thought and many more genes may be expressed in the infected cell. Herpesviruses express the enzymes necessary for viral DNA synthesis allowing herpesviruses to infect both dividing and quiescent cells. The HHVs have six blocks of orthologous genes with interspersed species-specific viral genes. There are substantial differences in their genomic sequences particularly in the unique coding regions of each herpesvirus. Antigenic analysis of both conserved and nonconserved genes is an important means for differentiation among herpesviruses despite some cross-reactions (eg, between HSV-1 and HSV-2).

Based on certain virologic similarities, the herpesviruses may be divided into three subfamilies α, β, and γ herpesviruses. HSV-1 and HSV-2, as well as VZV, are in the α subfamily, characterized by relatively rapid replication time and neuronal latency; CMV, HHV-6, and HHV-7 are in the β subfamily, characterized by slow replication rates and extremely limited host range; EBV and KSHV (HHV-8) are in the γ subfamily characterized by relatively rapid replication, replication in lymphocytes, and restricted host range. These characterizations are now made on the basis of genomic sequences but the original classifications have held up in the genomic era.

Cell tropism for the individual viruses varies significantly. HSV has the widest range; it can infect many different animal hosts and replicates in numerous animal and human host cells, although in nature it is only found in humans. VZV infects only humans and is best grown in cells of human origin, although some laboratory-adapted strains can grow in primate cell lines. Human CMV replicates well only in limited human cell lines including human foreskin fibroblasts. HHV-6 and HHV-7 preferentially grow in T-lymphocyte cell cultures. EBV does not replicate in most commonly used cell culture systems, but can be grown in continuous human or primate lymphoblastoid cell cultures where it is present in the latent state. KSHV infects many cell types but generally establishes latency in cultured cells, where only a low percentage of the cells support active replication.

Replication

The replication of HSV has been comprehensively studied and is representative of all herpesviruses, as shown in Figure 14–2. HSV generally causes lytic infection in epithelial cells and subsequently establishes latency in neuronal cells. The glycoproteins in the HSV envelope interact with cellular receptors, including initial binding to heparan sulfate and subsequent interaction with higher affinity receptors, leading to fusion with the cell membrane. For most herpesviruses fusion occurs at the cytoplasmic membrane but for some viruses or in specific cell types, the virus is first endocytosed and fusion occurs in the endosome. Fusion delivers tegument proteins into the cytoplasm as well as the capsid containing viral DNA. The capsid migrates to the nucleus where the genome is then extruded into the nucleus. In the nucleus, the viral genome circularizes and viral gene expression can be initiated. Transcription of the large, complex genome is sequentially regulated in three distinct classes of mRNAs: (1) immediate early (IE) mRNAs, also known as α genes, are synthesized 2 to 4 hours after infection. IE genes do not require de novo viral protein synthesis prior to expression and generally encode for proteins involved in regulation of viral gene expression and host defense; (2) early (E) mRNAs, or β genes, require prior protein synthesis of IE genes and generally encode proteins involved in viral replication (DNA binding proteins, DNA polymerase, thymidine kinase, etc), and (3) late (L) mRNAs, or γ genes, require viral genome replication for full expression and encode major structural proteins: capsid subunits, tegument proteins, and envelope glycoproteins. The early (E) proteins thymidine kinase and DNA polymerase are distinct from host cell enzymes and are, therefore, important targets of antiviral chemotherapy as discussed later. Synthesis of IE genes is required for E genes and E genes shut off the IE genes. The E genes are required for viral genomic replication, which in turn is required for optimal synthesis of most L genes. However, some of the late structural proteins are produced to lower levels independently of genome replication. Viral DNA replication occurs in a rolling circle fashion producing high–molecular-weight DNA concatemers. Genomic concatemers are cleaved and packaged into preassembled capsids in the nucleus.

Herpesvirsues assemble in the nuclei and a proteolytic cleavage event is necessary for the maturation of the capsid. A viral protease is responsible for the maturation. The envelope is acquired from the inner lamella of the nuclear membrane. Budding occurs at the nuclear membranes, and virions are then transported through the ER and Golgi. Re-envelopment and de-envelopment through the ER and Golgi and ultimately the cytoplasmic membrane is thought to occur. Host cell protein synthesis shut-off occurs for both α- and γ-herpesviruses and is thought to occur by cleavage of mRNAs by viral protein complexes. Ultimately, viral replication and host cell shut-off leads to death of the infected cell. Due to their long replication cycle, β-herpesviruses do not exhibit host cell shut-off.

Latency

In vivo, herpesviruses generally produce an initial lytic infection which is eventually controlled by the host immune system. However, during the initial infection, latent infection is also established. Latent infection allows all herpesvirus infection to be maintained for the life of the host. During latency, the genome of the virus is present in cells, but infectious virus is not recovered. The viral DNA is maintained as an episome in the nucleus. Integration is extremely rare. Latent infection is different from chronic infection in that the viral genome is not rapidly replicated and virions are not produced. During latency there is minimal viral gene expression with only 1 to 10 latent genes being regularly expressed, depending on the virus. Latent genes encode functions for maintenance of the viral episome, preventing host cell death and inhibiting the host immune response. Many herpesviruses also express microRNAs during latency. MicroRNAs are small regulatory RNAs that control gene expression without producing a peptide product. This allows the virus to alter host and viral gene expression without producing antigens that could be recognized by the host immune system. HSV-1 expresses only miRNAs during latent infection and no proteins, minimizing the ability of the immune system to recognize the latently infected cells. Periodic reactivation provides a constant source of new infections in the population. There is a range of reactivation rates depending on the virus and the host. In immunosuppressed patients, reactivation is more common and severe, indicating that the immune system must plays a role in the suppression of reactivation.

VIROLOGY

The genomes of herpes simplex 1 and 2 (HSV-1 and HSV-2, respectively) are both approximately 150 kbp of DNA. Although they are distinct epidemiologic and antigenic viruses, their genomes contain approximately 50% homology, making them the most closely related HHVs. Nearly all of the genes of HSV-1 have colinear homologs in HSV-2. HSV-1 and HSV-2 share many glycoprotein and structural antigens, but differences in glycoprotein B, among other glycoproteins, enable them to be distinguished antigenically. The viruses can also be distinguished by PCR assays as well.

HERPES SIMPLEX DISEASE

EPIDEMIOLOGY

HSV-1 is more often associated with disease “above the waist” or facial herpes, whereas HSV-2 is most often associated with genital infections or “below the waist” infections. However, an increasing number of genital infections are caused by HSV-1. HSV-1 is most often spread by direct contact of mucosal tissue, especially the lip area. Both HSV-1 and HSV-2 are prevalent worldwide. There are no known animal vectors for HSV-1 or HSV-2. Seroepidemiologic studies indicate that the prevalence of HSV antibody varies by age and socioeconomic status of the population studied. In most developing countries, up to 90% of the population has HSV-1 antibody by the age of 30 years. In the United States, HSV-1 antibody is found in 18% to 35% of children by the age of 5 years with the percentages varying according to the population studied. In the United States, the seroprevalence rises to approximately 60% to 70% by the age of 30 years for middle-class populations; among lower socioeconomic groups, however, the percentage is higher. Detection of HSV-2 antibody before puberty is less common. Direct sexual transmission is the major mode of spread. Approximately 15% to 30% of sexually active adults in Western industrialized countries have HSV-2 antibody and seropositive rates are positively correlated with the number of sexual partners. The virus can be isolated from the cervix and urethra of approximately 5% to 12% of adults attending sexually transmitted disease clinics; many of these patients are asymptomatic or have small, unnoticed lesions on penile or vulvar skin. Asymptomatic shedding accounts for transmission from a partner who has no active genital lesions and often no history of genital herpes. Genital herpes is not a reportable disease in the United States, but it is estimated that more than 1 million new cases occur per year.

PATHOGENESIS

Acute Infections

Both HSV-1 and HSV-2 initially infect and replicate in the mucoepithelial cells and initiate lytic or productive infection at the site of contact. Pathologic changes during acute infections consist of development of multinucleated giant cells (Figure 14–3), ballooning degeneration of epithelial cells, focal necrosis, eosinophilic intranuclear inclusion bodies, and an inflammatory response characterized by an initial polymorphonuclear neutrophil (PMN) infiltrate and a subsequent mononuclear cell infiltrate. The virus spreads to local sensory neurons and travels in retrograde fashion to the sensory ganglia that innervate the site of infection. In the case of facial herpes, the virus infects neurons in the trigeminal ganglia and in the case of genital herpes, the dorsal root or sacral ganglia. Latency is established in the ganglionic neurons. A round of replication may occur in the ganglia, but is not necessary for the establishment of latency.

Latent Infection

In humans, latent infection by HSV-1 has been demonstrated in trigeminal, superior cervical, and vagal nerve ganglia, and occasionally in the S2-S3 dorsal sensory nerve root ganglia. Latent HSV-2 infection has been demonstrated in the sacral (S2-S3) region. Latent infection of neurons by HSV does not result in the death of the cell. Multiple viral genomes exist in a circular extrachromosomal form in the nucleus, and transcription of only a small portion of the viral genome occurs, limited to a single viral transcript, the latency-associated transcript (LAT). The LAT encodes a number of miRNAs that serve as regulatory RNAs that can alter host cell gene expression without expressing foreign proteins. Because latency is established in nondividing neurons, HSV does not encode direct functions to maintain the viral episome. Latent infection does not require synthesis of early or late viral polypeptides and, therefore, antiviral drugs directed at the thymidine kinase enzymes or viral DNA polymerase do not eradicate the virus in its latent state.

A subset of patients exhibit overt clinical disease from reactivation of the virus. This can occur over the entire life of the host. However, people without obvious clinical disease can also reactivate and spread the virus through subclinical shedding. The mechanisms by which latent infection is reactivated are unknown. Precipitating factors that are known to initiate reactivation of HSV and subsequent clinical disease include exposure to ultraviolet light, sunlight, fever, excitement, emotional stress, and trauma (eg, oral intubation). However, it is clear that reactivation and viral shedding between overt disease episodes is common, and may account for some of the spread of the virus. Upon reactivation, the virus initiates lytic replication and virus particles travel down the neuronal axons, most often to a site near the site of initial infection. The epithelium is subsequently infected and leads to localized spread and ulceration in a subset of reactivations.

IMMUNITY

Host factors have a major effect on clinical manifestations of HSV infection. Many episodes of HSV infection are either asymptomatic or mildly symptomatic. Initial symptomatic clinical episodes of the disease are often more severe than recurrent episodes, likely due to the presence of anti-HSV antibodies and immune lymphocytes in persons with recurrent infections. Prior infection with HSV-1 may provide some level of protection against or shorten the duration of symptoms and lesions from subsequent infection with HSV-2 as a result of some degree of cross-protection, though dual infections certainly occur.

Both cellular and humoral immune responses are important in immunity to HSV. Neutralizing antibodies directed against HSV envelope glycoproteins appear to be important in preventing exogenous reinfection. Antibody-dependent cellular cytotoxicity (ADCC) may be important in limiting early spread of HSV. By the second week after infection, cytotoxic T lymphocytes can be detected, which have the ability to destroy HSV-infected cells before completion of the replication cycle. Conversely, in immunosuppressed patients, especially those with depressed cell-mediated immunity, reactivation of HSV may be associated with prolonged viral excretion and persistence of lesions as well as more disseminated lesions. During latency, the HSV-1 and HSV-2 do not express viral proteins and are thus effectively hidden from the immune system. However, the immune system plays a role in keeping latency in check as immunosuppression leads to more common reactivation. It is possible that the virus may initiate reactivation more often than previously thought and that the adaptive immune system shuts down those cells once they reactivate.

HSVs express a number of genes that have evolved to inhibit innate and adaptive immunity. There are a number of genes capable of inhibiting interferon pathways at different stages. HSV-1 also encodes an IE protein that blocks peptide loading onto MHC-I and prevents the complex from reaching the cell surface. Additionally, HSV inhibits apoptosis during both latent and lytic phases.

CLINICAL ASPECTS

MANIFESTATIONS

Herpes Simplex Type 1

Infection with HSV-1 is more often, associated with facial disease though it causes an increasing number of genital infections. It consists characteristically of grouped or single vesicular lesions that become pustular and coalesce to form single or multiple ulcers. On dry surfaces, these ulcers scab before healing; on mucosal surfaces, they reepithelialize directly. HSV can be isolated from almost all ulcerative lesions, but the titer of virus decreases as the lesions evolve. Infections generally involve ectoderm (skin, mouth, conjunctiva, and the nervous system).

Primary infection with HSV-1 is most often asymptomatic. When symptomatic, typically in children, it appears most frequently as gingivostomatitis, with fever and ulcerative lesions involving the buccal mucosa, tongue, gums, and pharynx. The lesions are painful, and the acute illness usually lasts 5 to 12 days. During this initial infection, HSV spreads to the sensory neurons and becomes latent within neurons of the trigeminal ganglia, the ganglia that innervated the oral and nasal area.

Lesions usually recur on a specific area of the lip and the immediate adjacent skin; these lesions are referred to as mucocutaneous and are commonly called “cold sores” or “fever blisters” (Figure 14–4). Lesions are typically unilateral. Their recurrence may be signaled by premonitory tingling or burning in the area. Systemic complaints are unusual, and the episode generally lasts approximately 7 days. It should be noted that HSV may be reactivated and excreted into the saliva with no apparent mucosal lesions present. HSV has been isolated from saliva in 5% to 8% of children and 1% to 2% of adults who were asymptomatic at the time.

In rare instances, HSV infects the finger or nail area. This infection, termed herpetic whitlow, usually results from the inoculation of infected secretions through a small cut in the skin or from needle sticks. Painful vesicular lesions of the finger develop and pustulate; they are often mistaken for bacterial infection and mistreated accordingly.

HSV infection of the eye is one of the most common causes of corneal damage and blindness in the developed world. Infections usually involve the conjunctiva and cornea, and characteristic dendritic ulcerations are produced. With recurrence of disease, there may be deeper involvement with corneal scarring. Occasionally, there may be extension into deeper structures of the eye, especially when topical steroids are used.

In rare cases, encephalitis may result from HSV-1 infection. Most cases occur in adults with high levels of anti-HSV-1 antibody, suggesting reactivation of latent virus in the trigeminal nerve root ganglion and extension of productive (lytic) infection into the temporoparietal area of the brain. Primary HSV infection with neurotropic spread of the virus from peripheral sites up the olfactory bulb into the brain may also result in parenchymal brain infection. Classically, HSV encephalitis affects one temporal lobe, leading to focal neurologic signs and cerebral edema. If untreated, mortality rate is approximately 70%. Clinically, the disease can resemble brain abscess, tumor, or intracerebral hemorrhage. Rapid diagnosis by polymerase chain reaction (PCR) of cerebrospinal fluid (CSF) has replaced brain biopsy as the diagnostic test. Intravenous acyclovir reduces the morbidity and mortality of the disease, especially if treatment is initiated early. There are small number of familiar genetic mutations leading to increased herpes encephalitis. These mutations appear to be in genes involved in specific innate immune responses.

Herpes Simplex Type 2

Genital herpes is a significant sexually transmitted disease. Both HSV-1 and HSV-2 can cause genital disease, and the symptoms and signs of acute infection are similar for both viruses. Seventy percent of the first episodes of genital HSV infection in the United States are caused by HSV-2, and genital HSV-2 disease is also more likely to recur than genital HSV-1 infection. Ninety percent of the HSV-2 antibody-positive patients have never had a clinically evident genital HSV episode. In many instances, the first clinical episode is years after primary infection.

Primary Genital Herpes Infection

For individuals who develop clinically evident primary genital HSV disease, the mean incubation period from sexual contact to onset of lesions is 5 days. Lesions begin as small erythematous papules, which soon form vesicles and then pustules (Figure 14–5). Within 3 to 5 days, the vesiculopustular lesions break to form painful coalesced ulcers that subsequently dry; some form crusts and heal without scarring. With primary disease, the genital lesions are usually multiple (mean number 20), bilateral, and extensive. The urethra and cervix are also infected frequently, with discrete or coalesced ulcers on the exocervix. Bilateral enlarged tender inguinal lymph nodes are usually present and may persist for weeks to months. About one-third of patients show systemic symptoms such as fever, malaise, and myalgia, and approximately 1% develop aseptic meningitis with neck rigidity and severe headache. First episodes of disease last an average of 12 days.

Recurrent Genital Herpes Infection

In contrast to primary infection, recurrent genital herpes is a disease of shorter duration, usually localized in the genital region and without systemic symptoms. A common symptom is prodromal paresthesias in the perineum, genitalia, or buttocks that occur 12 to 24 hours before the appearance of lesions. Recurrent genital herpes usually presents with grouped vesicular lesions in the external genital region. Local symptoms such as pain and itching are mild, lasting 4 to 5 days, and lesions usually last 2 to 5 days.

At least 80% of patients with primary, symptomatic, genital HSV-2 infection develop recurrent episodes of genital herpes within 12 months. In patients whose lesions recur, the median number of recurrences is four or five per year. They are not evenly spaced, and some patients experience a succession of monthly attacks followed by a period of quiescence. Over time, the number of recurrences decreases by a median of one-half to one recurrence per year. Recurrences result from reactivation of virus from dorsal root ganglia. Recurrent infections due to reinfection with a different strain of HSV-2 are extremely rare. Recurrent viral shedding from the genital tract often occurs without clinically evident disease.

Neonatal Herpes

Neonatal herpes usually results from transmission of virus during delivery through infected genital secretions from the mother. In utero infection, though possible, is uncommon. In most cases, severe neonatal herpes is associated with primary infection of a seronegative woman near the time of delivery. This results in an intense viral exposure of a seronegative infant as it passes through the birth canal. The incidence of symptomatic neonatal herpes simplex infection varies greatly among populations, but it is estimated at between 1 per 6000 and 1 per 20 000 live births in the United States. Because a normal immune response is absent in the neonate born to a mother with recent primary infection, neonatal HSV infection is an extremely severe disease with an overall mortality rate of approximately 60%, and neurologic sequelae are high in those who survive. Manifestations vary. Some infants show disseminated vesicular lesions with widespread internal organ involvement and necrosis of the liver and adrenal glands; others have involvement of the central nervous system only, with listlessness and seizures.

DIAGNOSIS

Herpes simplex viruses (HSVs) can be cultured in cell lines inoculated with infected secretions or lesions. The cytopathic effects of HSV can usually be demonstrated 24 to 48 hours after inoculation of the culture. Isolates of HSV-1 and HSV-2 can be differentiated by staining virus-infected cells with type-specific monoclonal antibodies. A direct smear prepared from the base of a suspected lesion and stained by either Giemsa or Papanicolaou method may show intranuclear inclusions or multinucleated giant cells typical of herpes (Tzanck test), but this is less sensitive than viral culture and not specific. Similar changes can be seen in cells infected with VZV. Enzyme immunoassays and immunofluorescence are rapid and relatively sensitive assays for direct detection of herpes antigen in lesions. Although early versions of these noncultural tests lacked sensitivity, more recent procedures have correlations with culture that approach 90%. Serology should not be used to diagnose active HSV infections, such as those affecting the genital or central nervous systems; frequently there is no change in antibody titer when reactivation occurs. Serology can be useful in detecting those with asymptomatic HSV-2 infection. PCR of CSF and blood is the best test to diagnose HSV encephalitis.

TREATMENT

Several antiviral drugs that inhibit HSV have been developed. The most commonly used is the nucleoside analog acyclovir, which is converted by a viral enzyme (thymidine kinase) to a monophosphate form and then by cellular enzymes to the triphosphate form. The triphosphate form is then incorporated by the viral polymerase into the ongoing replicating viral genome leading to chain termination due to the lack of a hydroxyl group to build upon. Acyclovir significantly decreases the duration of primary infection and has a lesser but definite effect on recurrent mucocutaneous HSV infections. If taken daily, it has been shown to suppress recurrences of genital and oral–labial HSV. In its intravenous form, it is effective in reducing mortality of HSV encephalitis and neonatal herpes. Acyclovir-resistant HSV has been recovered from immunocompromised patients with persistent lesions, especially those with acquired immunodeficiency syndrome (AIDS). Foscarnet is active against acyclovir-resistant HSV.

The US Food and Drug Administration has approved both valacyclovir and famciclovir for the treatment of recurrent genital HSV. Valacyclovir is an oral prodrug of acyclovir with better bioavailability than acyclovir (54% compared with 15-20%). It is rapidly converted to acyclovir and, in every characteristic except absorption, it is identical with the parent compound. Valacyclovir is not more effective than acyclovir, but can be given in lower doses and less frequently (500 mg twice daily). Famciclovir is the prodrug of another guanosine nucleoside analog, penciclovir. The bioavailability of famciclovir is also high (77%). After conversion, penciclovir must be phosphorylated, similarly to acyclovir. Penciclovir has a longer tissue half-life than acyclovir and can be given as 125 mg twice daily for treatment of recurrent genital HSV. Valacyclovir and famciclovir are now also approved for chronic suppression of recurrent genital HSV. Valacyclovir taken daily was shown to decrease spread between discordant partners in a long-term study.

PREVENTION

Avoiding contact with individuals with lesions reduces the risk of spread; however, virus may be shed asymptomatically and transmitted from the saliva, urethra, and cervix by individuals with no evident lesions. Safe sexual practices should reduce transmission. Acyclovir has been shown to reduce asymptomatic shedding and transmission of genital herpes, especially from males to females. Because of the high morbidity and mortality rates of neonatal infection, special attention must be paid to preventing transmission during delivery. Where active HSV lesions are present on maternal tissues, Cesarean section delivery may be used to minimize contact of the infant with infected maternal genital secretions, but Cesarean delivery may not be effective if rupture of the membranes precedes delivery by more than several hours. Avoiding the birth canal is particularly important if the mother has a primary HSV infection late during pregnancy. There is no current HSV vaccine available though a number have been under study for years.

VIROLOGY

Varicella-zoster virus (VZV) has the same general structural and morphologic features of herpes simplex and other HHVs, but it contains distinct glycoproteins and is antigenically different. The genome of VZV is approximately 125 kbp, which is the smallest genome of the HHVs. Similar to HSV, VZV encodes a thymidine kinase and is responsive to acyclovir. Cellular features of infected cells such as multinucleated giant cells and intranuclear eosinophilic inclusion bodies are similar to those of HSV. VZV is more difficult to isolate in cell culture than HSV. The virus often remains attached to the membrane of the host cell with less release of virions into fluids and thus does not spread well in culture. However, this is not the case in vivo, where it is the most infectious human herpesvirus.

VARICELLA-ZOSTER DISEASE

EPIDEMIOLOGY

VZV infection is ubiquitous. In temperate climates, greater than 90% of people contract varicella (chickenpox) by the time they reach adulthood, and most cases occur before the age of 10 years. In contrast, the mean age at infection in tropical countries is over 20 years, and the seroprevalence at the age of 70 years may be only 50%. The virus is highly contagious, with attack rates among susceptible contacts of 75% making it the most infectious of the HHVs. Varicella occurs most frequently during the winter and spring months. The incubation period is 11 to 21 days. The major mode of transmission is respiratory route, although direct contact with vesicular or pustular lesions may result in transmission. Communicability is greatest 24 to 48 hours before the onset of rash and lasts 3 to 4 days into the rash phase. Virus is difficult to isolate from patients once lesions have crusted over.

Zoster or shingles results from a reactivation of VZV and occurs in approximately 20% of the population. Zoster occurrences depend on the severity of the initial infection. Zoster is rare in children. After the age of 50 years the incidence of Zoster increases dramatically. Zoster provides a constant source of VZV for spread. Initial infection with VZV or spread from zoster will result in Varicella.

PATHOGENESIS

Spread of virus by the respiratory route leads to infection of the patient’s upper respiratory tract followed by replication in regional lymph nodes and primary viremia. The latter results in infection of the reticuloendothelial system and a subsequent secondary viremia associated with T lymphocytes. After secondary viremia, there is infection of the skin and ultimately, a host immune response. Latency is established in the same sensory ganglia as HSV-1 and HSV-2.

The relation between zoster and varicella was first described by Von Bokay in 1892, when he observed several instances of varicella in households after the introduction of a case of zoster. On the basis of these epidemiologic observations, he proposed that zoster and varicella were different clinical manifestations of a single agent. The cultivation of VZV in vitro by Weller in 1954 confirmed Von Bokay hypothesis that the viruses isolated from chickenpox and from zoster (or shingles) are identical. Latency of VZV occurs in sensory ganglia, primarily in the trigeminal ganglia, but the dorsal root ganglia of adults can also support latent infection. Herpes zoster (shingles) occurs when latent VZV reactivates and multiplies within a sensory ganglion and then travels back down the sensory nerve to the skin. In immunocompetent patients, the rash of herpes zoster is generally confined to the area of the skin (ie, dermatome) innervated by the sensory ganglion in which reactivation occurs.

IMMUNITY

Both humoral immunity and cell-mediated immunity are important factors in the VZV immune response. Cell-mediated immunity is thought to be important for cessation of spread of VZV in the body as most spread is cell associated. Reinfection with VZV is rare and is prevented by circulating antibody, whereas reactivation of VZV is apparently controlled by cell-mediated immunity.

The increase in the incidence and severity of herpes zoster observed with increasing age in immunocompetent individuals is correlated with an age-related decrease in VZV-specific cellular immunity. Beginning in the fifth decade of life, there is a marked decline in cellular immunity to VZV, which can be measured by delayed cutaneous hypersensitivity as well as by a variety of in vitro assays. This occurs many years before any generalized decline in cellular immunity. In patients with depressed cell-mediated immune responses, especially those with bone marrow transplants, Hodgkin disease, AIDS, and lymphoproliferative disorders, reactivation often occurs, and is more frequent and severe.

CLINICAL ASPECTS

MANIFESTATIONS

Varicella (Chickenpox)

VZV produces a primary infection in normal children characterized by a generalized vesicular rash termed chickenpox or varicella. After clinical infection resolves, the virus persists for decades with no clinical manifestation. Chickenpox lesions generally appear on the back of the head and ears, and then spread centrifugally to the face, neck, trunk, and proximal extremities. Involvement of mucous membranes is common, and fever may occur early in the course of disease. Lesions appear in stages of spread during the course of disease (Figure 14–6); this characteristic was one of the major features used to differentiate varicella from smallpox, in which lesions are concentrated on the extremities and all have a similar appearance. Skin lesions form rapidly as fluid-filled vesicles that become turbid after 1 to 2 days and then crust over. Varicella lesions are pruritic (itchy), and the number of lesions may vary from 10 to several hundred.

Immunocompromised children may develop progressive varicella, which is associated with prolonged viremia and visceral dissemination as well as pneumonia, encephalitis, hepatitis, and nephritis. Progressive varicella has an estimated mortality rate of 20%. In thrombocytopenic patients, the lesions may be hemorrhagic. Susceptible adults upon primary infection have a higher risk (15 times) for VZV pneumonia during chickenpox. Mortality in children aged 1 to 14 years is less than 1 in 100 000 patients. However, mortality increases in primary infection of adult populations to 25 per 100 000 patients between 30 and 49 years of age.

Herpes Zoster (Shingles)

Reactivation of VZV is associated with the disease herpes zoster (shingles). Although zoster can be found in patients of all ages, the frequency of patients developing shingles greatly increases with advancing age. Clinically, pain in a sensory nerve distribution may herald the onset of the eruption, which occurs several days to 1 or 2 weeks later. The vesicular eruption is usually unilateral, involving one to three dermatomes (Figure 14–7). New lesions may appear over the first 5 to 7 days. Multiple attacks of VZV reactivation and disease are uncommon; if recurrent attacks of a vesicular eruption occur in one area of the body, HSV infection should be considered.

The complications of VZV infection are varied and depend on age and host immune factors. Postherpetic neuralgia is a common complication of herpes zoster in elderly adults. It is characterized by persistence of pain in the dermatome for months to years after resolution of the lesions of zoster and appears to result from damage to the involved nerve root. Immunosuppressed patients may develop localized zoster followed by dissemination of virus with visceral infection, which resembles progressive varicella. Bacterial superinfection is also possible. Maternal varicella infection during early pregnancy can result in fetal embryopathy with skin scarring, limb hypoplasia, microcephaly, cataracts, chorioretinitis, and microphthalmia. Severe varicella can also occur in seronegative neonates, with a mortality rate as high as 30%.

DIAGNOSIS

Varicella or herpes zoster lesions can be diagnosed clinically as the rash is quite characteristic, and therefore laboratory diagnosis is not generally necessary. However, the lesions are occasionally difficult to distinguish from those caused by HSV. Scrapings of lesions may reveal multinucleated giant cells characteristic of herpesviruses, but cytologic examination does not distinguish HSV lesions from those due to VZV. For rapid viral diagnosis, varicella-zoster antigen can be identified in cells from lesions by immunofluorescent antibody staining. VZV can be isolated from vesicular fluid or cells inoculated onto human diploid fibroblasts. However, the virus is difficult to grow from zoster (shingles) lesions older than 5 days, and cytopathic effects are usually not seen for 5 to 9 days, therefore PCR detection is more commonly done. PCR of CSF may be useful in the diagnosis of VZV encephalitis; culture is rarely positive.

TREATMENT

Acyclovir has been shown to reduce fever and skin lesions in patients with varicella, and its use is recommended in healthy patients over 18 years of age. There are insufficient data to justify universal treatment of all healthy children and teenagers with varicella. In immunosuppressed patients, controlled trials of acyclovir have been effective in reducing dissemination, and the use of this agent is indicated. In addition, controlled trials of acyclovir have demonstrated effectiveness in the treatment of herpes zoster in immunocompromised patients. Acyclovir may be used to treat herpes zoster in immunocompetent adults, but it appears to have only a modest impact on the development of postherpetic neuralgia, an important complication of zoster. Treatment should be started within 3 days of the onset of zoster. VZV is less susceptible than HSV to acyclovir, so the dosage for treatment is substantially higher. Famciclovir or valacyclovir are more convenient and may be more effective.

PREVENTION

High-titer immune globulin (VariZig) administered within 96 hours of exposure is useful in preventing infection or ameliorating disease in patients at risk for severe primary infection (eg, immunosuppressed children with contact of patients with varicella or zoster). Once skin lesions have occurred, however, high-titer immune globulin has not proved useful in ameliorating disease or preventing dissemination. Immune globulin is not indicated for the treatment or prevention of reactivation (ie, zoster or shingles). In nonimmunosuppressed children, varicella is a relatively mild disease, and passive immunization is not indicated.

An attenuated live virus vaccine (Oka) developed in Japan is effective in both immunosuppressed and immunocompetent persons, and is now recommended for routine use in healthy children (Table 14–2). Routine immunization at 12 to 15 months with single or in a combination vaccine with MMR, and a second dose at 4 to 6 years of age is now recommended. For older seronegative people, two doses 4 to 8 weeks apart is recommended. In immunocompromised patients who are susceptible to varicella, chickenpox can be extremely serious, even fatal. In these patients, the live vaccine appears to be protective, although it is not approved for this use in the United States. The vaccine is used routinely in immunocompetent seronegative adults, especially those with occupational risk, such as healthcare workers. The vaccine can be helpful when given to a seronegative, immunocompetent adult shortly after exposure.

Vaccination for zoster is also possible. The zoster vaccine is a high dose version of the varicella vaccine. It is approved for all people over the age of 60 years, those most susceptible to VZV reactivation and zoster. The vaccine stimulates the waning cellular immunity, and thereby decreases reactivation. The zoster vaccine has been shown to be approximately 50% effective in preventing zoster and slightly more effective in eliminating postherpetic neuralgia. Chronic conditions such as renal failure, heart disease, or diabetes are not contraindications, but this vaccine is not recommended for immunosuppressed patients. Varicella is a highly contagious disease and rigid isolation precautions must be instituted in all hospitalized cases.

VIROLOGY

Human cytomegalovirus (CMV) is a β-herpesvirus named for the cytopathic effect it produces in cell culture. In addition to nuclear inclusions (“owl’s eye cells”), CMV produces perinuclear cytoplasmic inclusions and enlargement of the cell (cytomegaly) (Figure 14–8). CMV possesses the largest genome of the HHVs (approximately 240 kbp). Similar to the α-herpesviruses, CMV gene expression is highly regulated with the sequential appearance of IE, E, and L gene products, though the replication cycle is much slower. Based on genomic and phenotypic heterogeneity, innumerable strains of CMV exist. Antigenic variations have been observed, but are not of clinical importance. Laboratory-adapted strains grown in cell culture rapidly lose a 10 to 15 kbp region of CMV genomic DNA that limits tropism in culture.

CYTOMEGALOVIRUS DISEASE

EPIDEMIOLOGY

CMV is ubiquitous. In developed countries approximately 50% to 70% of adults have developed antibody with even higher percentages in lower socioeconomic strata and in the developing world. Age-specific prevalence rates show that approximately 10% to 15% of children are infected by CMV during the first 5 years of life, after which the rate of new infections levels off. The rate subsequently increases by 1% to 2% per year during adulthood. Infection probably occurs through close personal contact, including sexual contact with a virus-excreting person. CMV has been isolated from saliva, cervical secretions, semen, urine, and white blood cells for months to years after infection. Excretion of CMV is especially prolonged after congenital and perinatal infections, with 35% of infected infants excreting virus for as long as 5 years after birth. Transmission of infection in day care centers has been shown to occur from asymptomatic excreters to other children and, in turn, to seronegative parents. By 18 months, up to 80% of infants in day care centers are infected and actively excreting virus in saliva and urine. Seroconversion rates in seronegative parents who have children attending day care centers are approximately 20% per year. In contrast to day care centers, there is no substantial evidence for spread of CMV infection to healthcare workers in hospitals.

Latent infection occurs in leukocytes and their precursors and accounts for transfusion transmission, but this route is relatively infrequent—only 1% to 2% of blood units are believed to be infectious. Organ donation may also transmit latent virus, which causes primary infection in CMV-seronegative recipients and reinfection in seropositive patients.

PATHOGENESIS

CMV infects vascular endothelial cells and leukocytes and produces characteristic inclusions in the former. In vitro, CMV DNA can be demonstrated in monocytes showing no cytopathology, indicating a restricted growth potential in these cells. It is conjectured that these as well as the CD34+ pluripotent stem cells that can differentiate into monocytes maintain latency for CMV.

CMV can cause disease by a variety of different mechanisms, including direct tissue damage and immunologic damage. Although direct infection and damage of mucosal epithelial cells in the lung is a potential mechanism for pneumonia, animal models have suggested that immunologic destruction of the lung by the host immune response to CMV infection may be the major mechanism of viral disease in this tissue. This hypothesis is supported by the observation that the degree of viral infection in lung tissue cannot account for the severity of CMV pneumonia; likewise, the disease does not respond well to antiviral therapy. Although cytolytic T-lymphocyte activity may contribute to lung pathology, cytokines released by these cells have also been implicated.

IMMUNITY

Both humoral and cellular immune responses are important in CMV infections. In immunocompetent persons, clinical disease, if it occurs at all, results from primary infection. Reactivation and viral excretion in cervical excretions or semen is invariably subclinical. In immunocompromised patients, both primary infection and reactivation are much more likely to be symptomatic. Furthermore, CMV infection of monocytes results in dysfunction of these phagocytes in immunocompromised patients, which may increase predisposition to fungal and bacterial superinfection. When latently infected monocytes are in contact with activated T lymphocytes, the former are activated to differentiate into macrophages that produce infectious virus. These monocyte–T cell interactions may occur after transfusion or transplantation and may explain, not only transmission of CMV, but also activation of latent virus in the allograft recipient.

CLINICAL ASPECTS

MANIFESTATIONS

CMV can be transmitted vertically to the fetus in utero leading to deafness or other congenital defects. Worldwide, 1% of infants excrete CMV in urine or nasopharynx at delivery as a result of infection in utero. On physical examination, 90% of these infants appear normal or asymptomatic; however, long-term follow-up has indicated that 10% to 20% go on to develop sensory nerve hearing loss, psychomotor mental retardation, or both. Infants with symptomatic illness (about 0.1% of all births) have a variety of congenital defects or other disorders, such as hepatosplenomegaly, jaundice, anemia, thrombocytopenia, low birth weight, microcephaly, and chorioretinitis. Almost all infants with clinically evident congenital CMV infection are born to mothers who experienced primary CMV infection during pregnancy. The apparent explanation is that the fetus is exposed to virus in the absence of maternal antibody. It is estimated that one-third of maternal primary infections are transmitted to the fetus and that fetal damage is most likely to occur in the first trimester. Congenital infection frequently also results from reactivation in the mother with spread to the fetus, but such infection rarely leads to congenital abnormalities because the mother also transmits antibody to the fetus. It is more common for second children to have congenital CMV infection. This is thought to be due to the first child obtaining CMV in day care, a common place for spread, and infecting the naïve pregnant mother.

In contrast to the devastating findings with some congenital infections, neonatal infection acquired during or shortly after birth is rarely associated with adverse outcomes. Most population-based studies have indicated that 10% to 15% of all mothers are excreting CMV from the cervix at delivery. Approximately one-third to one-half of all infants born to these mothers acquire infection. Illness is rare in perinatally infected infants unless the infant is premature or immunocompromised. CMV can also be efficiently transmitted from mother to child by breast milk, but these postpartum infections are also usually benign.

As with intrapartum acquisition of infection, most CMV infections during childhood are asymptomatic. By contrast, in healthy young adults and adults, CMV may cause a mononucleosis-like syndrome. In immunosuppressed patients, both primary infection and reactivation may be severe. For example, in patients receiving bone marrow transplants, interstitial pneumonia caused by CMV is a leading cause of death (50-90% mortality rate), and in patients with AIDS, CMV often disseminates to visceral organs, causing chorioretinitis, gastroenteritis, and neurologic disorders. CMV retinitis is the main cause of blindness in patients with AIDS.

DIAGNOSIS

Laboratory diagnosis of CMV infection depends on (1) detecting CMV cytopathology, antigen, or DNA in infected tissues; (2) detecting viral DNA or antigen in body fluids; (3) isolating the virus from tissue or secretions; or (4) demonstrating seroconversion. CMV can be grown in serially propagated diploid fibroblast cell lines. Demonstration of viral growth generally requires 1 to 14 days, depending on the concentration of virus in the specimen and whether fluorescence immuno-staining is used to speed detection. The presence of large inclusion-bearing cells in urine sediment may be detected in widespread CMV infection. This technique is insensitive, however, and provides positive results only when large quantities of virus are present in the urine. Culture of blood to detect viremia is now superseded by detection and quantitation of CMV antigen in peripheral blood leukocytes or detection of CMV DNA in plasma or leukocytes by PCR. These procedures are significantly more sensitive than culture.

Because of the high prevalence of asymptomatic carriers and the known tendency of CMV to persist weeks or months in infected individuals, it is frequently difficult to associate a specific disease entity with the isolation of the virus from a peripheral site. Thus, the isolation of CMV from urine of immunosuppressed patients with interstitial pneumonia does not constitute evidence of CMV as the cause of that illness. CMV pneumonia or gastrointestinal disease is best diagnosed by demonstrating CMV inclusions in biopsy tissue.

The procedures listed below are recommended to facilitate the diagnosis of CMV infection in specific clinical settings:

1. Congenital infection—Virus culture or viral DNA assay positive at birth or within 1 to 2 weeks (to distinguish from natally or perinatally infected infants, who will not begin to excrete virus until 3-4 weeks after delivery).

2. Perinatal infection—Culture-negative specimens at birth but positive specimens at 4 weeks or more after birth suggest natal or early postnatal acquisition. Seronegative infants may acquire CMV from exogenous sources, such as from blood transfusion.

3. CMV mononucleosis in nonimmunocompromised patients—Seroconversion and presence of IgM antibody specific for CMV are best indicators of primary infection. Urine culture positivity supports the diagnosis of CMV infection, but may reflect remote infection because positivity may continue for months to years. A positive blood assay for CMV antigen or DNA, however, is diagnostic in this patient population.

4. Immunocompromised patients—Demonstration of virus by viral antigen or DNA in blood documents viremia. Demonstration of inclusions or viral antigen in diseased tissue (eg, lung, esophagus, or colon) establishes the presence of CMV infection, but does not provide proof that CMV is the cause of disease unless other pathogens are excluded. Seroconversion is diagnostic but rarely occurs, especially in patients with AIDS, because more than 95% of these patients are seropositive for CMV before infection with human immunodeficiency virus (HIV). CMV-specific IgM antibody may not be present in immunocompromised transplant patients, especially during reactivation of virus. Conversely, in patients with AIDS, this antibody frequently is present even when clinically important infection is absent.

TREATMENT

Ganciclovir, a nucleoside analog of guanosine structurally similar to acyclovir, has been shown to inhibit CMV replication, prevent CMV disease in patients with AIDS and transplant recipients, and reduce the severity of some CMV syndromes such as retinitis and gastrointestinal disease. Combining immune globulin with ganciclovir appears to reduce the very high mortality from CMV pneumonia in bone marrow transplant recipients more than that achieved with ganciclovir alone. However, unlike acyclovir, ganciclovir has some toxicity. Foscarnet, a second approved drug for therapy of CMV disease, is also efficacious. Its toxic effects are primarily renal, whereas ganciclovir is most apt to inhibit bone marrow function. Ganciclovir is phosphorylated by the viral kinase, UL97 and acts as a chain terminator when incorporated by the CMV DNA polymerase. Valganciclovir, also approved for use as a CMV therapeutic, is a prodrug of ganciclovir and provides increased bioavailability. Foscarnet inhibits the CMV polymerase and is used as a second-line drug for CMV. A third drug, cidofovir, a nucleotide analog, is approved for therapy of retinitis, but its use is limited to ganciclovir-resistant infections in immunosuppressed patients because of nephrotoxicity.

PREVENTION

The use of blood from CMV-seronegative donors or blood that is treated to remove white cells decreases transfusion-associated CMV. Similarly, the disease can be avoided in seronegative transplant recipients by using organs from CMV-seronegative donors. Safe sexual practices including condom usage may reduce transmission. There is currently no vaccine available.

VIROLOGY

Epstein-Barr virus (EBV) is the main etiologic agent of infectious mononucleosis (IM), African BL and nasopharyngeal carcinoma (NPC) and is associated with a number of other B-cell lymphomas. EBV is a γ-herpesvirus and its DNA genome is 172 kbp in length. In vivo, EBV has tropism for both human B lymphocytes and epithelial cells. In vitro, EBV can be cultured only in human or some primate B cells as well as limited epithelial cultures. In cultured B lymphocytes, the virus establishes a latent infection. Therefore, the virus does not produce cytopathic effects or the characteristic intranuclear inclusions of other herpesvirus infections. A low percentage of cultured primary human B cells infected with EBV grow out to form immortal lymphoblastoid cell lines (LCLs) that can grow permanently in culture and maintain EBV infection. The viral DNA in LCLs remains in a circular extrachromosomal, nonintegrated form, and is only very rarely found in the integrated state. Lytic replication can be found in LCLs induced to reactivate by cross-linking the B-cell receptor or other chemical means and follows similar gene regulation cascades as the other herpesviruses.

EBV Latency

A number of different forms of latency have been described for EBV, each with a different viral gene profile. Three main types of latent infection have been characterized, though slightly different gene expression has been described in specific settings. LCLs support type III latency which is characterized by the expression of four EBV nuclear antigens (EBNAs), including EBNA-1 that is necessary to maintain the episome, and two integral membrane proteins, LMP-1 and LMP-2. A number of small RNAs are also expressed including the abundant EBERs and the BARTs that encode a number of regulatory miRNAs. Type II latency, found in NPC cells, does not express the full range of EBNA proteins but expresses many of the other viral genes found in type III latency. Type I latency is found in most BL cells and has more limited gene expression with only EBNA-1 and small regulatory RNAs expressed.

EPSTEIN-BARR VIRUS DISEASE

EPIDEMIOLOGY

Over 90% of the population is seropositive for EBV worldwide. In developing countries, most children are infected by the age of 2 years, whereas in the developed world EBV infection occurs more often in late childhood or adolescence. When primary infection with EBV is delayed until the second decade of life or later, it is accompanied by symptoms of IM in about 50% of the cases. There are two main strains of EBV (types 1 and 2) that both circulate widely, and can coinfect a single individual. EBV is spread by direct contact of oropharyngeal secretions. The virus can be routinely cultured from saliva in 10% to 20% of healthy adults and is intermittently recovered from most seropositive individuals. It is of low contagiousness, and most cases of IM are contracted after repeated contact between susceptible persons and those asymptomatically shedding the virus. Secondary attack rates of IM are low (<10%), because most family or household contacts already have antibody to the agent. IM has also been transmitted by blood transfusions; most transfusion-associated mononucleosis syndromes, however, are attributable to CMV.

PATHOGENESIS

Although EBV initially infects epithelial cells in the oral environment, the hallmark of EBV disease involves subsequent infection of B lymphocytes and polyclonal B-lymphocyte activation with benign proliferation. The virus enters B lymphocytes by means of envelope glycoprotein binding to a surface receptor CR2 or CD21, which is the receptor for the C3d component of complement system; 18 to 24 hours later, EBNAs are detectable within the nucleus of infected cells. Infection is associated with immortalization and proliferation of the B cell. The EBV-infected B lymphocytes are polyclonally activated to produce immunoglobulin and express a lymphocyte-encoded membrane antigen that is the target of host cellular immune responses to EBV-infected B lymphocytes. During the acute phase of IM, up to 20% of circulating B lymphocytes demonstrate EBV antigens. After infection subsides, EBV can be isolated from only about 1% of such cells.

EBV has been associated with several lymphoproliferative diseases, including African BL and posttransplant lymphomas in immunocompromised patients. EBV is also associated with an epithelial tumor, NPC. The factors that render the EBV infections oncogenic in these cases are not clear, but a few of the type III latency genes are necessary for immortalization of B cells, in particular latent membrane protein-1. The distribution of EBV infections in Africa has suggested an infectious cofactor, such as malaria, which may lead to further activation of infected B cells and enable BL formation. In vivo, EBV-associated lymphomas have been shown to be of both monoclonal and polyclonal origin. In BL, translocations, involving the c-myc oncogene and immunoglobulin heavy or light loci, are almost invariable. These translocations lead to increased c-myc expression and subsequent expression of oncogenic pathways that may contribute to B-cell activation and ultimately to malignancy. EBV is present in many forms of NPC and is thought to play an etiologic role. However, environmental carcinogens and genetic factors may also be operative. Some breakdown in immune surveillance also appears to play a role in the development of malignancy, because immunosuppressed patients are more prone to develop EBV-associated B-cell lymphomas. Recent studies suggest an association of EBV with Hodgkin lymphoma in young adults.

IMMUNITY

Virus-induced IM is associated with circulating antibodies against specific viral antigens, as well as against unrelated antigens found in sheep, horse, and some bovine red blood cells. The latter, referred to as heterophile antibodies, are a heterogeneous group of predominantly IgM antibodies long known to correlate with episodes of IM, and are commonly used as diagnostic tests for the disease. They do not cross-react with antibodies specific for EBV, and there is no good correlation between the heterophile antibody titer and the severity of illness. Cutaneous anergy and decreased cellular immune responses to mitogens and antigens are seen early in the course of mononucleosis. The “atypical” lymphocytosis associated with IM is caused by an increase in the number of circulating T cells, which appear to be activated cells developed in response to the virus-infected B lymphocytes. With recovery from illness, the atypical lymphocytosis gradually resolves, and cell-mediated immune functions return to preinfection levels, although memory T cells maintain the capacity to limit proliferation of EBV-infected B cells. In rare cases, the initial EBV-induced proliferation of B cells is not contained, and EBV lymphoproliferative disease ensues. This syndrome is most often seen in immunocompromised organ transplant recipients.

CLINICAL ASPECTS

MANIFESTATIONS

Infectious Mononucleosis

Most primary EBV infections are asymptomatic. However, infections in the second decade of life often lead to clinically apparent IM. IM is characterized by fever, malaise, pharyngitis, tender lymphadenitis, and splenomegaly. These symptoms persist for days to weeks; they slowly resolve. Complications such as laryngeal obstruction, meningitis, encephalitis, hemolytic anemia, thrombocytopenia, or splenic rupture may occur in 1% to 5% of patients.

Lymphoproliferative Syndrome

Patients with primary or secondary immunodeficiency are susceptible to EBV-induced lymphoproliferative disease. The incidence of these lymphomas is 1% to 2% after renal transplantations and 5% to 9% after heart–lung transplantations. The risk is greatest in patients experiencing primary EBV infection rather than reactivation. The most characteristic symptoms are persistent fever, lymphadenopathy, and hepatosplenomegaly.

Burkitt Lymphoma

In sub-Saharan Africa, endemic Burkitt lymphoma (BL) is the most common malignancy in young children, with an incidence of 8 to 10 cases per 100 000 people per year. Endemic BL is almost always associated with EBV. The risk is greatest in equatorial Africa, where there is a high incidence of malaria. Endemic BL commonly occurs along the jawline or orbit of the eye in children. Endemic BL is thought to result from an early EBV infection that produces a large pool of infected B lymphocytes. Malarial infection may further increase the size of this pool and provide a constant antigenic challenge. Such stimuli can increase the chances of c-myc chromosomal translocations, which are pathognomonic for this lymphoma. Serologic screening for increased IgA antibody levels to both VCA and early EBV antigens can be used for early diagnostic purposes. In the United States and other countries where BL is not endemic, EBV is associated with 15% to 30% of sporadic BL while c-myc translocations are still invariable in all forms.

Other EBV-associated lymphomas

• Hodgkin lymphoma: There are four histologic subtypes of Hodgkin lymphoma (HL) with a broad range of EBV association. Overall, approximately 30% of HL are associated with EBV in the United States and other developed countries. A much higher percentage of HL is associated with EBV in developing countries. In HL EBV is present in Reed-Sternberg cells, large often multinucleated cells of B-cell origin.

• Non-Hodgkin lymphoma: Additionally EBV is associated with a lower percentage of other B-cell lymphoma including diffuse large B-cell lymphoma (DBCL). EBV is also associated with some T-cell lymphomas as well.

Nasopharyngeal Carcinoma

Nasopharyngeal carcinoma (NPC), an epithelial derived tumor, is endemic in southern China, where it is responsible for approximately 25% of the mortality from cancer. The high incidence of NPC among the southern Chinese people suggests that in addition to EBV, genetic or environmental factors may also be important in the pathogenesis of the disease.

AIDS Patients

In patients with AIDS, several distinct additional EBV-associated diseases may occur, including hairy leukoplakia of the tongue, interstitial lymphocytic pneumonia (especially in infants), and increased incidence of lymphoma described earlier. EBV is also associated with posttransplant lymphoproliferative disorders (PTLD).

DIAGNOSIS

Laboratory analysis of EBV IM is usually documented by the demonstration of atypical lymphocytes and heterophile antibodies, or positive EBV-specific serologic findings. Hematologic examination reveals a markedly raised lymphocyte and monocyte count with more than 10% atypical lymphocytes, called Downey cells (Figure 14–9). Atypical lymphocytes, though not specific for EBV, are present with the onset of symptoms and disappear with resolution of disease. Alterations in liver function tests may also occur, and enlargement of the liver and spleen is a common finding.

Though not specific for EBV, tests for heterophile antibodies are used most commonly for diagnosis of IM. In commercial kits, animal erythrocytes are used in simple slide agglutination methods, which incorporate absorptions to remove cross-reacting antibodies that may develop in other illnesses, such as serum sickness. The IM heterophile antibody is absorbed by sheep erythrocytes, but not by guinea pig kidney cells. Heterophile antibodies can usually be demonstrated by the end of the first week of illness, but are occasionally delayed until the third or fourth week. They may persist many months.

Approximately 5% to 15% of EBV-induced cases of IM in adults and a much greater proportion in young children and infants fail to induce detectable levels of heterophile antibodies. In these cases, the EBV-specific serologic tests summarized in Table 14–3 may be used to establish the diagnosis. The panel to be tested includes antibodies to VCA, which rise quickly and persist for life. Antibodies to EBNAs rise later in disease (after about 1 month) and also persist in low titers for life. Thus, a high titer to VCA and no titer to EBNA suggest recent EBV infection, whereas antibody titers to both antigens are indicative of past infection. The presence of IgM antibody to VCA is theoretically diagnostic of acute, primary EBV infection, but low levels may occur during reactivation of EBV, and cross-reactions with antigens of other herpesviruses occur. Persistent antibody to early antigens (anti-EA, -D, or -R) may be correlated with severe disease, NPC (anti-EA-D), or African BL (anti-EA-R), but are not useful in diagnosing IM. Isolation of EBV from clinical specimens is not practical, because it requires fresh human B cells or fetal lymphocytes obtained from cord blood.

TREATMENT AND PREVENTION

Treatment of IM is largely supportive. More than 95% of patients recover uneventfully. In a small percentage of patients, splenic rupture may occur; restriction of contact sports or heavy lifting during the acute illness is recommended. Lytic replication of EBV has been shown to be sensitive to acyclovir, and acyclovir can decrease the amount of replication of EBV in tissue culture and in vivo. Despite this antiviral activity, systemic acyclovir makes little or no impact on the clinical illness. Laryngeal obstruction should be treated with corticosteroids. Hairy leukoplakia in patients with AIDS does respond to acyclovir treatment. There is currently no approved vaccine for Epstein-Barr virus and few preventative measures other than limiting saliva transmission.

In 1986, a herpesvirus, now called human herpesvirus type 6 (HHV-6), was identified in cultures of peripheral blood lymphocytes from patients with lymphoproliferative diseases. HHV-6 is β-herpesvirus subfamily. The virus morphologically similar to other herpesviruses with similar replication patterns of other herpesviruses. HHV-6 replicates in lymphoid tissue, especially CD4+ T lymphocytes, and has two distinct variants, A and B, that are genetically disparate enough that some consider them different species.

EPIDEMIOLOGY

Of the herpesviruses, HHV-6 is the most rapidly spread and is shed in the throats of 10% of babies by age 5 months, 70% by 12 months, and 30% of adults. Greater than 90% of the population has antibody to this virus by the age of 5 years.

MANIFESTATIONS

HHV-6 type B is the main etiologic agent of exanthem subitum (roseola), and both types A and B can cause acute febrile illnesses with or without seizures or rashes. Exanthem subitum generally occurs in infants aged 6 months to 1 year. In the first 6 months, infants are generally protected by the mother’s IgG. Exanthem subitum is characterized by fever (usually about 39°C) for 3 days, followed by a faint maculopapular rash spreading from the trunk to the extremities, which begins during defervescence. Exanthem subitum is one of the six classic childhood exanthems.

HHV-6 also appears to reactivate in transplant recipients. It may contribute to graft rejection and clinical illnesses such as meningoencephalitis, pneumonia, and bone marrow suppression after bone marrow transplantation. The virus reactivates in other immunocompromised patients including those with AIDS, lymphoma, and leukemia, but its clinical significance is not known. Attempts have been made to associate HHV-6 persistence with many other disease states including multiple sclerosis and chronic fatigue syndrome. However, due to the ubiquitous nature of the virus, disease association is difficult to assess.

HHV-6 infects mainly T lymphocytes and establishes a latent infection in T cells, but may be activated to a productive lytic infection by mitogenic stimulation. Resting lymphocytes and lymphocytes from normal immune individuals are resistant to HHV-6 infection. In vivo, HHV-6 replication is controlled by cell-mediated factors.

DIAGNOSIS

Primary virus infection can be documented by seroconversion. Active virus infection can be documented by culture, antigenemia, or DNA detection in the blood (by PCR). Because asymptomatic viremic reactivation is common, it is very difficult to use these tools to identify HHV-6 as the cause of febrile or other miscellaneous syndromes.

TREATMENT

Definitive therapy has not been established, but like the better characterized β-herpesvirus, CMV, HHV-6 appears to be susceptible in vitro to ganciclovir and foscarnet. It is less susceptible to acyclovir, because the virus has no thymidine kinase.

Isolation of human herpesvirus 7 (HHV-7) was first reported in 1990. The virus was isolated from activated CD4+ T lymphocytes of a healthy individual. The CD4 molecule appears to be a receptor for virus attachment. HHV-7 is closely related to HHV-6 and is in the β-herpesvirus genus. Seroepidemiologic studies indicate that this virus usually does not infect children until after infancy, but that nearly 90% of children are antibody positive by 3 years of age. As with HHV-6, this virus is frequently isolated from saliva, and close personal contact is the probable means of transmission. There is little disease associated with HHV-7, however, it may also be a cause of exanthem subitum, but the association has only been found in rare cases. The diagnosis of acute infection can be made by the demonstration of seroconversion. No treatment has been identified.

During the AIDS epidemic in the 1980, in the United States, Kaposi Sarcoma (KS) occurred in 20% to 30% of gay or bisexual males with AIDS but in only around 1% of hemophiliacs with AIDS. This led to the proposal that there was another infectious agent associated with KS. In 1994, unique viral DNA sequences were identified in KS tumors using subtractive hybridization analysis. The sequences bore homology to γ-herpesviruses and were used to clone the entire 165 kbp genome of the eighth HHV, commonly known as KS-associated herpesvirus (KSHV) or HHV-8. KSHV is found in 100% of KS tumors.

EPIDEMIOLOGY

KSHV is the least widespread HHV. In the United States, 5% or less of healthy blood donors are seropositive. Worldwide the seroprevalence varies dramatically. In central Africa, where KS is endemic, KSHV seroprevalence can reach 50%. Classic KS is more common in Southern Italy where seroprevalence approaches 25%, whereas in Northern Italy where KS is less common the seroprevalence is closer to 10%. As noted earlier, in the United States, KS was common in the gay and bisexual AIDS community where the seroprevalence rates perfectly match the KS rates at around 25%, whereas in hemophiliacs with AIDS the seroprevalence rates were similar to healthy blood donors. The relationship of seroprevalence rates and KS probability were critical for collaring KSHV as the etiologic agent of KS. KSHV is also associated with two rare lymphoproliferative diseases, primary effusion lymphoma, where KSHV is nearly 100% associated, and multicentric Castleman disease (MCD), where it is associated with 50% of AIDS-related cases.

KSHV seropositive rates correlate with numbers of sexual partners and was originally thought to be transmitted sexually. However, the virus is not found in sexual secretions but is shed in saliva. Because the virus is not ubiquitous like the other saliva-transmitted herpesvirus, it is not likely to be easily transmitted by kissing and may require more prolonged intimate contact.

PATHOGENESIS

KSHV infects the oral epithelium and can be shed into saliva for transmission. KSHV is also found in the B-cell fraction of peripheral blood mononucleocytes. In B cells, KSHV is predominantly in the latent state although lytic antigens can be found in a low percentage of the cells. In KS tumors, KSHV is found in the main tumor cell, the spindle cell, a cell of endothelial origin. KSHV is found in all spindle cells in later stage tumors. Again, the virus is found predominantly in the latent state, though 1% to 5% of the spindle cells support lytic antigens and likely replication and virus production. In culture, KSHV can infect many cell types where it establishes latency in most of the cells. Similar to the KS tumor, a low percentage of endothelial cells infected in culture also express lytic antigens.

CLINICAL MANIFESTATIONS

KS—Four Forms of Disease

There are four main forms of KS: classic, endemic, iatrogenic, and epidemic or AIDS associated. The forms vary in the degree of severity but are indistinguishable at the pathologic level. KSHV is associated with all four forms.

1. Classic KS—Originally described in the 1800s by Moriz Kaposi, it is a rare, fairly indolent tumor mainly found on the lower extremities. It is mostly seen in elderly men of Mediterranean origin and was also described in Ashkenazi jews.

2. Endemic KS—In the middle of the 20th century, KS became common in central Africa, where in countries like Uganda it is the most common tumor reported in hospitals. It is more aggressive than classic KS and tumors can be seen higher on the extremities and in the oral cavity and the torso.

3. Iatrogenic KS—KS also arises in posttransplant patients, but generally regresses upon the removal of immunosuppression.

4. Epidemic or AIDS-associated KS—This is the most aggressive form of KS, with the tumors often appearing first in the mouth, on the torso, and face, and can also be found on internal organs. Without treatment for HIV, it can lead to death.

Primary effusion lymphoma (PEL): PELs have high mortality and KSHV is associated with nearly 100% of this pleural cavity B-cell lymphoproliferative disease. EBV is also present in 50% to 70% of PELs and may play a contributing role in these cases. Unlike Burkitt lymphoma, there are no known obvious genetic abnormalities in PELs.

Multicentric Castleman disease (MCD): MCD is a B-cell lymphoproliferative disease of the lymph nodes and 50% of AIDS-associated MCD is associated with KSHV. The infected cells in MCD have a much higher percentage of cells expressing lytic antigen than other KSHV-associated tumors.

DIAGNOSIS

Diagnosis for KSHV infection is currently imperfect. Immunofluorescence with sera from infected patients is a standard technique but has a sensitivity of only 70% to 90%. PCR from peripheral blood mononucleocytes of patients with KS is possible, but in seropositive patients without KS, KSHV DNA is difficult to detect.

TREATMENT AND PREVENTION

A number of antiherpesviral drugs inhibit lytic replication of KSHV with foscarnet being the most active followed by ganciclovir. There is evidence that treatment with ganciclovir is positively indicated for MCD because it is a more lytic disease. No treatment for latently infected cells or vaccine is available.