Prevention and control of herpesvirus diseases

ii,eD O lilt public~~health. tMostUofate senies epubcesdllrbique.L A art~~~~~iceswil beowritten, byth cesta artiLes auoint douncssentubla inv opresentposibtonk n e se- tigonsu tuemle pans les o- Xups ate X / ~~~~lected fields, and, over a / maine conside'r. Des experts // / / ~~~~~~~~periodof years, will cover /couvriront ainsi successive-/ / / / ~~~~~~~~manydifferent aspects of /ment de nombreux aspects des / r ,. + X X ~~~~~the biomedical sciences and /sciences biom6dicales et de la/ e.opinl public health. Most of the /sante publique. La plupart de / / / / ~~~~~~~~articles will be written, by /ces articles auront donc ete / / / ~~~~~~~~invitation, by acknowledged /reddige's sur demande par les / experts on the subject. specialistes les plus autorises. Bulletin ofthe WorldHealth Organization, 63 (2): 185- 201 (1985) © World Health Organization 1985 Prevention and control of herpesvirus diseases* Part 1. Clinical and laboratory diagnosis and chemotherapy A WHO MEETING1 The herpesvirus diseases are increasing in importance as a public health problem throughout the world. Members of the human herpesvirus family are global in distribution and infect 60-95% of the world's population, both in developed and in developing countries. Illnesses associated with herpesviral infections vary from simple blisters to deadly encephalitis. In numerical terms, primary cytomegalovirus infection is a more common cause of congenitally acquired disease than primary rubella and results in severe handicap. The apparent increasing incidence ofgenital herpes, which may induceperinatal infections associated with a high rise in perinatal morbidity and mortality, is a cause of concern. Neonatally acquired infection has a high mortality and many of the survivors have permanent sequelae. Herpes simplex virus, cytomegalovirus and varicella-zoster virus reactivations frequently lead to death in patients undergoing immunosuppressive therapy. Close association between herpes simplex infection and cervical neoplasia has been reported. Unlike other viruses, the herpesviruses usually remain latent throughout life. Viral reactivations, induced by various factors (hormonal, environmental, physical or chemical agents), may result in a large variety of illnesses, with high prevalences in both developing and developed countries. Advances are currently being made in diagnostic, preventive and therapeutic approaches to the human herpesviruses. There are new methods for rapid disease diagnosis and for epidemiological investigations, including the rapid analysis of viruses and antibodies. New antiherpetic drugs have been licensed in some countries, and vaccines are undergoing trials. This report summarizes the information currently available and makes recommendations to enhanceprogress in the control ofdiseases caused by the herpes- viruses. * This article is based on the report of an Informal WHO Meeting on Recent Progress towards the Prevention and Control of Herpesvirus Diseases, which was held in Geneva on 14-18 November 1983. The next issue of the Bulletin will include the second and final part, together with the full list of references. A French translation of this article will appear in a later issue of the Bulletin. Requests for reprints should be sent to Chief, Virus Diseases, World Health Organization, 1211 Geneva 27, Switzerland. ' The names of the participants will appear with Part 2. 4514 -185 A WHO MEETING The herpes group of viruses is attracting increasing attention among both the lay and medical communities because of the increasing awareness of their importance as a public health problem in causing disease throughout the world. The changing epidemiological features of these infections are leading to new problems, e.g., the emergence of cytomegalovirus as a major cause of fetal infections producing congenital malformations, and the increasing prevalence of primary genital infections with herpes simplex virus. Infections with these viruses have also appeared as major complications in immunosuppressed patients receiving organ transplants. A close association between herpes simplex infection and cervical neoplasia has been reported. The fact that herpesviruses can become latent in the host is a major problem in the control of these diseases. This virus group is thus a major cause of morbidity and mortality. At the same time, new tools are becoming available to the investigator for the study and control of this group of viruses. There are new methods for rapid detection and diagnosis and for epidemiological investigations including the rapid analysis of viruses and antibodies. Drugs are now becoming available for the systemic treatment of some members of the herpesvirus group. The seriousness of the diseases caused by herpesviruses makes it desirable to prevent them. New knowledge from molecular genetics and the use of monoclonal antibodies have opened up fresh possibilities for vaccine development. The aim of such vaccine studies should be to prevent primary herpesvirus infections and associated diseases. THE HUMAN HERPESVIRUS FAMILY In the past 50 years, more than 80 distinct herpesviruses have been isolated from a wide variety of animal species and been grouped in the family Herpesviridae on the basis of common structural features. The virus particles are composed of a DNA-containing core surrounded by (a) a protein capsid, (b) a protein coat designated as tegument, and (c) an envelope. The envelope is essential for infectivity. Glycoproteins in the viral envelope are responsible for attachment of the virus particle to the cell, for the penetration of the virus into the cell, and for eliciting neutralizing antibodies. Even though the various herpes- viruses cannot be differentiated by electron microscopy, few virus families exhibit as much variation as the members of the Herpesviridae family. These viruses are, however, readily differentiated on the basis of their biological properties, immunological specificity of their virions, and the size, base composition, and arrangement of their genomes. Common and important herpesviruses of humans include herpes simplex virus types 1 and 2, varicella-zoster virus, Epstein-Barr virus, and cytomegalovirus. The common names Table 1. Human herpesvirus Common name/Alternative name' ICTV nameb Subfamily Herpes simplex virus type 1 (HSV-1 )/Human (alpha) herpesvirus 1 Simplexvirus 1 Alphaherpesvirinae Herpes simplex virus type 2 (HSV-2)/Human (alpha) herpesvirus 2 Simplexvirus 2 Alphaherpesvirinae Varicella-zoster virus (VZV)/Human (alpha) herpesvirus 3 -C Alphaherpesvirinae Cytomegalovirus (CMV)/Human (beta) herpesvirus 5 _d Betaherpesvirinae Epstein-Barr Virus (EBV)/Human (gamma) herpesvirus 4 Lymphocryptovirus Gammaherpesvirinae a From International nomenclature of diseases, Vol. 2: Infectious diseases, Pt 3: Viral diseases. Geneva, CIOMS, 1983, pp. 8-20. b ICTV: International Committee for the Taxonomy of Viruses. c No approved name so far. d Proposed name (not approved): Poikilovirus. 186 PREVENTION AND CONTROL OF HERPESVIRUS DISEASES 187 Table 2. Main characteristics of the human Herpesviridaea Properties of the virus particles 1. Nucleic acid: DNA, double-stranded, linear 32-75 G + C mole %, relative molecular mass approximately 100-1 50 x 106. 2. Proteins: more than 20 structural polypeptides with relative molecular mass from 12 000 to 220 000. 3. Lipid: exact proportion in the total weight is unknown, probably variable; located in the virion envelope. 4. Carbohydrate: exact proportion in the total weight is unknown; identified largely as covalently linked to envelope proteins. 5. Physico-chemical characteristics: buoyant density (CsCI) of virion 1.20-1.29 g/cm3; relative molecular mass 109 or higher. 6. Morphology. The virion (1 20-200 nm in diameter) consists of four structural components. The core consists of a fibrillar spool on which the DNA is wrapped. The ends of the fibres are anchored to the underside of the capsid shell. The capsid (100-1 10 nm) is an icosahedron with 5 capsomeres on each edge; it contains 1 50 hexameric and 12 pentameric capsomeres. The hexameric capsomeres contain a hole running halfway through the long axis. The tegument surrounding the capsid consists of globular material that is often distributed asymmetrically and may be variable in amount. The envelope, a bilayered membrane surrounding the tegument, has surface projections. The intact envelope is impermeable to negative stain. 7. Antigenic properties: neutralizing antibody reacts with major viral glycoproteins located in the viral envelope; an Fc receptor for immunoglobulin G may be present in the virion envelope. 8. Effects of virus suspensions on cells: fusion and agglutination occur rarely or only under very special conditions in the absence of replication. Virus replication 1. Entry: the viral envelope adsorbs to receptors on the plasma membrane of the host cell, ultimately fuses with the membrane, and releases the capsid into the cytoplasm; a DNA-protein complex is then translocated into the nucleus. 2. Replication: viral DNA is transcribed in the nucleus; messenger RNAs generated from the transcripts are translated in the cytoplasm; viral DNA is replicated in the nucleus and is spooled into preformed, immature nucleocapsids. 3. Maturation and egress: the ability to infect cells is acquired as capsids become enveloped by budding through the inner lamelIa of the nuclear membrane and, in some instances, through other membranes of the cell; virus particles accumulate in the space between the inner and outer lamellae of the nuclear membrane and in cysternae of the endoplasmic reticulum; virus particles are released by transport to the cell surface through the modified endoplasmic reticulum. a Modified from B. Roizman (1). and scientific designations of human herpesviruses are given in Table 1, and their properties are listed in Table 2. These viruses have been grouped according to their growth properties both in vivo and in vitro. Herpes simplex virus types 1 and 2 (HSV-1 and 2), which share common antigenic deter- minants, and varicella-zoster virus (VZV) have been placed in the subfamily of Alpha- herpesvirinae. Upon infection with members of this subfamily, e.g., HSV-1 and HSV-2, cytopathic effects spread rapidly in cell culture resulting in mass destruction of susceptible cells. Both epithelial and fibroblastic cells from various species readily support replication of these viruses in vitro and a variety of experimental models exist for in vivo study. Cytomegalovirus (CMV) is a member of the subfamily Betaherpesvirinae. Members of this subfamily characteristically induce cell enlargement (cytomegalia) and formation of inclusion bodies in the nuclei and cytoplasm of cells following both in vitro and in vivo infection. In culture, the propagation of cytopathic effects is very slow and the virus grows only in human fibroblasts. The host range is restricted to humans, and no experimental animal model exists for CMV. In vivo, it is not certain what type of cell is the target of infection although cytomegalic inclusions can be found in many organs. Epstein-Barr virus (EBV) belongs to the Gammaherpesvirinae subfamily, which is composed of lymphotropic viruses.a Infection of lymphocytes with EBV is frequently a EBV has been discussed at length at meetings convened by the International Agency for Research on Cancer (IARC). The results have been published by IARC, e.g., in DE-THt, G. & ITo,Y., ed., Nasopharyngeal carcinoma: etiology and control. Lyon, International Agency for Research on Cancer, 1978. A WHO MEETING arrested either at a prelytic stage, with persistence and minimum expression of the viral genome, or at a lytic stage, with resultant cell death but without production of complete virions. In vitro viral growth occurs only in B lymphocytes. In vivo, the experimental host range includes some species of marmosets. Herpesviruses usually remain latent for the lifetime of the host following primary infection, but the cell type harbouring the latent infection varies depending on the virus subfamily. Herpes simplex viruses are readily found latent in the dorsal root and auto- nomic ganglia. The cells harbouring latent members of the Betaherpesvirinae have not been identified, but recipients of transplant organs and blood products from antibody-positive donors may become infected. Gammaherpesvirinae are readily recovered from lymphocytes and the nasopharynx. Transmission of human herpesviruses occurs principally by contact between mucosal surfaces (HSV-1, HSV-2, CMV, EBV). All of them may traverse the placental barrier but this is particularly true for CMV. Other means of transmission are organ transplants, and, in the case of CMV, the use of infected blood products and breast milk. Finally, some (like VZV) are transmitted by airborne routes. Herpesviruses have been associated with malignant diseases in humans: for example, there is a very strong association between Epstein-Barr virus and Burkitt's lymphoma in African children and between this virus and nasopharyngeal carcinoma. Herpes simplex type 2 virus has frequently been associated with cervical and vulvar carcinoma, but the etio- logical significance of this association remains unclear. A considerable amount of basic research is being done in many countries throughout the world. Information is accumulating and aiding our understanding of (1) the pathogenesis of the diseases caused by herpesviruses, (2) the mechanisms by which cellular and humoral immunity curtail virus infections, (3) the antigens that elicit protective immunity in the human host, and (4) the mechanisms by which drugs suppress virus multiplication. Definitions The following definitions are used in this report: Serotype: viruses of the same serotype share most of their nucleotide sequences and anti- genic determinants although they may exhibit antigenic and genetic variability; only one serotype is recognized for CMV, VZV, and EBV, and two serotypes for HSV (HSV-l and HSV-2). Primary infection: a first infection of a human host with any serotype of a virus. First infection: the first successful exposure of an individual to a given herpesvirus sero- type, irrespective of an infection by other serotypes. First clinical episode: the first clinically recognized disease; it may, but need not be, the primary infection. Secondary clinical episodes: these are recognized disease episodes occurring in indivi- duals with previous clinical episode(s). Latent infection (latency): the presence in a particular tissue of virus in a non-infectious form, which can be induced to replicate by specific stimuli, e.g., by the culture of tissue in vitro. Reactivation: the induction of replication of latent virus; this may, or may not, lead to clinical lesions. Asymptomatic shedding: viral excretion in the absence of detectable clinical symptoms; 188 PREVENTION AND CONTROL OF HERPESVIRUS DISEASES this may be the consequence of persistent virus multiplication or a reactivation of latent virus. Exogenous (super) infection: infection of a previously infected individual with a virus of the same serotype from an exterior source; exogenous virus can be differentiated from reactivated virus only by laboratory techniques. Recurrent infection: presence of virus at body surfaces or in secretions as a result of reactivation of latent virus, or as a consequence of reinfection with virus from an external source. Recurrent lesions: lesions containing virus derived from reactivation within the dermatome or at or near the portal of entry of the virus into the body; this definition applies to HSV and VZV. Recurrent disease: severe clinical manifestations which may follow virus replication in the host. Except in the case of HSV, a rise in antibody titres may be the sole manifestation of recurrence. CLINICAL SPECTRUM The herpesviruses are ubiquitous agents that produce protean clinical manifestations. Although these infections are generally clinically inapparent or mild, they may result in severe disease which is occasionally fatal (2). The following clinical manifestations are associated with herpesviruses: (a) HSV-from simple blisters to deadly encephalitis; (b) CMV-from mild disease in adults to severe mental retardation and other handicaps following congenital infection; (c) VZV-from mild disease in the normal population to deadly varicella in immuno- compromised patients. Unlike other viruses, the herpesviruses usually remain latent throughout life. Reactivated virus may cause a variety of illnesses; among others these include: (a) HSV-keratoconjunctivitis, recurrent herpes labialis and genitalis, and rarely encephalitis; (b) CMV -pneumonitis in immunocompromised patients; (c) VZV-herpes-zoster, and rarely disseminated varicella with encephalitis. With the exception of varicella, a large proportion of primary infections are asymptomatic or produce minimal clinical consequences. However, under certain conditions (old age, immunodeficiency (whether congenital or iatrogenic), a different site of entry of the virus, large size of the inoculum, etc.), primary or first infections with symptoms are more likely to occur. Clinical manifestations may also result from reactivation of a latent virus but, with the exception of encephalitis, they are of lesser severity. In immunocompromised patients, recurrences of all herpesviruses may cause serious disease and even death. Herpes simplex virus Primary infection. Primary HSV infection is symptomatic in 10-50Oo of subjects. Its typical clinical presentations are gingivostomatitis, keratoconjunctivitis, and vesicular 189 A WHO MEETING Table 3. Clinical spectrum of primary infections due to human herpesviruses Cytomegalovirus Herpes simplex viruses Varicella-zoster virus Epstein-Barr virus Typical clinical presentation Mononucleosis Skin vesicles Chickenpox Mononucleosis mucosal ulcerations Less frequent manifesta- tions Pneumonitis Urethritis Pneumonitis Pneumonitis Hepatitis Tender adenopathies Meningitis Encephalitis Encephalitis Meningitis Encephalitis Myelitis Haemolytic anaemia Encephalitis Myelitis Meningitis Retinitis Dysfunction of Arthritis Cerebellar ataxia Vasculitis - skin rash autonomic system Reye's syndrome Bell's palsy Guillain-Barr6 syndrome Keratoconjunctivitis Bell's palsy Vasculitis rash Thrombocytopenia Disseminated rash Guillain-Barr6 syndrome Arthritis Rupture of spleen Pancytopenia Table 4. Characteristics of genital herpes" First episode Recurrent Primary Primary Non-primary HSV-1 HSV-2 HSV-2 HSV-2 (n = 20) (n = 189) (n = 76) (n = 362) Systemic symptoms (%) 58 62 16 10 Meningitis (%) 16 26 1 - Mean number of lesions 24.3 15.5 9.5 6.1 Mean duration of lesions (days) 22.7 18.6 15.5 9.9 Mean duration of shedding (days) 11.1 11.4 6.8 4.1 Mean duration of local pain (days) 12.5 11.8 8.7 4.9 Extragenital lesions (%) 10 18 8 4 Shedding from cervix (%) 80 88 65 12 Duration of shedding from cervix (days) - 11.4 - 3.2 ' Modified from Corey et al. 13). mucocutaneous lesions, usually restricted to the oral and genitoanal areas (2, 3). They may be associated with systemic symptoms (Table 3). These symptoms tend to be more severe and more frequent in HSV-2 than in HSV-1 infections. The most severe but fortunately relatively rare manifestation is encephalitis, which is frequently fatal. Aseptic meningitis as a manifestation of first-episode HSV infection occurs more frequently in adults than in children. HSV-2 infections are generally less severe in those who were previously infected with HSV-1 (first non-primary infection) (Table 4). Recurrent infection. Secondary clinical HSV episodes produce local signs and symptoms that are milder and, with the exception of encephalitis, are less frequently associated with systemic clinical manifestations than first clinical episodes (2). However, recurrent ocular lesions may result in blindness. Subjects suffering from pathological skin conditions like atopic eczema or burns may develop Kaposi's varicelliform eruption, a generalized form of cutaneous herpes. The immunocompromised subjects are also at higher risk of developing 190 PREVENTION AND CONTROL OF HERPESVIRUS DISEASES generalized cutaneous lesions than normal individuals. However, HSV encephalitis has not been associated with immune deficiency. Perinatal infection. Intrauterine HSV infection is rare (2, 4). Perinatal infection (intra- partum) occurs in 1 out of 4000-30 000 deliveries (2, 4). These infections generally present symptoms and their clinical course may be either localized or disseminated. The latter is usually characterized by hepatitis, encephalitis, and pneumonitis with or without skin involvement. Localized infections may involve the CNS, eye, skin, or oral cavity. When untreated, the disease has a high mortality rate (65%). Survivors are frequently handi- capped for life. Cytomegalovirus Primary infection. Primary infections with CMV in immunologically competent hosts are generally asymptomatic (2). In normal adults, the most characteristic presentation for a first clinical episode of CMV infection is a mononucleosis-like syndrome which is usually mild. The syndrome is characterized by fever, malaise, and lymphocytosis with numerous atypical lymphocytes, but the characteristics of EBV mononucleosis (sore throat, generalized lymphadenopathy, and presence of heterophile antibodies) are lacking. Occasionally, primary CMV infection may be accompanied or followed by systemic mani- festations involving almost any organ system (Table 3). Patients with inherited immuno- deficiency diseases, or receiving immunosuppressive drugs, are at a significantly higher risk of a severe generalized infection and increased mortality (2, 5, 6). The clinical spectrum in transplant recipients is well defined. It is characterized by viral excretion, at single or multiple sites, rise in antibody, and the following signs and symptoms: fever for more than 3 days, pneumonitis without other causes, leukopenia (less than 3000 WBC), and hepatic dysfunction (in the absence of hepatitis B and atypical lymphocytosis). The onset is between 2 and 4 months after the transplantation, with a peak in the second month. The mean duration of the syndrome is 70 days. For reasons that are not clear, the main complication of CMV after bone marrow transplants is interstitial pneumonitis, which may occur in as many as 50% of patients and has a mortality rate ranging from 40% to 80% (6). Blood transfusion and CMV infection. The post-transfusion mononucleosis-like syndrome is a well known complication of surgery with transfusions. The syndrome develops between 3 and 7 weeks after the operation and is more severe in seronegative patients. In general, this complication is more likely to occur when fresh blood (less than 48 hours of storage) is used and the risk is directly proportional to the volume transfused. Leukocyte-poor blood is less likely to cause this syndrome. In adults it has been estimated that the risk is approximately 2.5/o per unit of fresh blood transfused. In recent years, transfusion-related CMV infection has also been described in newborn infants (7). The syndrome occurs primarily in premature infants with a birth weight of less than 1500 g. The clinical characteristics of the syndrome are presented in Table 5. The risk of acquiring CMV infection is directly related to the number of transfusions, and the number of seropositive donors. The risk of disease and mortality is significantly reduced by the presence of maternally derived (transplacental) IgG antibodies. In one important study, transfusion-acquired CMV infection in premature infants resulted in 40/o mortality, compared with a 6.6% mortality rate for uninfected patients of similar characteristics who were cared for at the same institution. It also resulted in a significant prolongation of hospitalization (7). Recurrent infection. Recurrent CMV infection in immunologically competent indivi- duals is almost always asymptomatic (but is expressed by viral excretion or antibody rises 191 A WHO MEETING or both). However, in sharp contrast, immunocompromised patients are at risk of severe generalized manifestations (2, 5, 6). In renal transplant recipients, these clinical manifestations are more serious but their duration is longer in primary than recurrent infections. Table 5. Characteristics of transfusion-acquired CMV in preterm infants ahb Percentage frequency Hepatosplenomegaly 93 Septic appearance 88 Congenital and perinatal CMV infec- Deteriorated respiration 71 tions. CMV is currently recognized as the Grey pallor 71 most common cause of congenital viral Fever (more than 38 OC) 36 infection in humans occurring in approxi- Death 20 mately 1% of all live births (4). Only about Atypical lymphocytosis (> 8%) 935%0 of infants with congenital CMV Thrombocytopenia (<75000 /mm3) 29 infection have typical cytomegalic inclusion disease, which is characterized by petechia a Sick preterm infants hospitalized for > 1 month. with thrombocytopenia, hepatospleno- b Modified from BALLARD, R. A. ET AL. Acquired cyto- megaly, jaundice with direct hyperbili- megalovirus infection in preterm infants. American journal rubinaemia and abnormal liver function of diseases of children, 133: 482-485 119791. tests, microcephaly, chorioretinitis, and typical lymphocytosis. Another 5% have less severe clinical involvement, and 9007o have no clinical manifestations at birth. Disease is more likely to occur with congenital infection resulting from primary as opposed to recurrent maternal infections. Cytomegalic inclusion disease may in over 90% of cases lead to sequelae, which include microcephaly, hydrocephalus, hearing loss, retinitis, and mental retardation. These delayed complications may also occur in infants with inapparent congenital infection at birth (approximately 100/o of cases) (4). CMV is also one of the most common agents transmitted from mother to baby during the first weeks and months of life, with an incidence ranging from 10% to 60% of infants. Transmission occurs as a result of exposure to maternal genital secretions at birth and to breast milk. In this respect, breast-feeding and the rate of seropositivity among nursing women are the two most important factors. Perinatally acquired CMV infections are generally asymptomatic (95% of cases). Occasionally pneumonitis develops at between 4 and 12 weeks of age, and more rarely hepatitis and haemolytic anaemia. Varicella infection Primary infection. Varicella virus is usually transmitted by the airborne route, and trans- mission by direct personal contact with an infectious patient accounts for only a small fraction of cases. The primary infection, after an incubation period of 3 weeks, always results in symptomatic disease which is characterized by fever, malaise, and typical papulo- vesicular skin lesions. Mucosal surfaces are generally involved. The acute disease is usually self-limiting and resolves in about 10 days, leaving behind lifelong immunity to reinfection. Other clinical manifestations are rare. Susceptible adults are at high risk for life- threatening varicella. Primary infection may lead to the development of less frequent but more severe clinical manifestations (see Table 3), of which pneumonitis can be a particular problem, and sometimes death ensues. Patients with inherited immunodeficiency diseases, or who are receiving immuno- suppressive drugs, are also at a significantly higher risk of severe generalized infections and death. In these patients, varicella is characterized by a severe skin eruption with either bullous formation or haemorrhage into the skin. There is often high fever and dissemi- nation of infection (in 3007o of cases) to various organs producing hepatitis, pneumonitis, 192 PREVENTION AND CONTROL OF HERPESVIRUS DISEASES pancreatitis, and encephalitis. Pneumonitis is particularly severe; it usually occurs between 3 and 7 days after the onset in 25% of cases and the mortality rate is approximately 7%. Recurrent infection. A reactivation of varicella-zoster virus leads to the appearance of herpes zoster along the sensory distribution of the involved dorsal spinal or cranial nerve roots. Systemic clinical manifestations are common (2). Aborted varicella-zoster reactivations may also occur and are characterized by exquisite pain without skin eruptions. Immunocompromised patients, particularly those under treatment for Hodgkin's disease and for bone marrow transplantation, are at higher risk (35-50Oo) of developing herpes zoster. These individuals may develop extensive skin lesions which may become necrotic. Generalized cutaneous dissemination of virus occurs in 30-50% of them and significant visceral involvement (e.g., hepatitis, pneumonitis, encephalitis) in approximately 10%. Bacterial superinfections are common. The mortality rate for immunosuppressed patients with disseminated herpes zoster is 3-5%. Congenital infection. Intrauterine infection with varicella, occurring during the first and second trimester of gestation, has been described only occasionally (4). This infection may cause fetal wastage and birth defects. Intrauterine infections occurring near term may cause typical varicella in the newborn with varying degrees of severity, depending on the transfer of maternal specific IgG antibody. Rarely, in children exposed to VZV in utero, herpes zoster develops at an early age. General management of patients Besides specific therapy, the management of patients with known active herpesvirus infections should also deal with measures to prevent further spread. The risk of themselves acquiring cytomegalovirus from infected patients also presents a problem to health care workers, especially those who may be pregnant. Recent studies, however, indicate that occupational contact with cases presents no greater risk than that faced by young women in the community at large (8). Strict handwashing and other general precautionary pro- cedures for dealing with potentially infected patients seem to provide sufficient protection from CMV infection. The problem ofCMV infections following blood transfusions can be avoided by the use of only seronegative blood, which has been successfully demonstrated in newborn infants (7). Preliminary data suggest that the use of glycerolized, frozen red blood cells is also a safe procedure. Varicella virus is known to be transmitted readily in the hospital setting. The risk is par- ticularly significant for immunocompromised patients or for susceptible health-care providers since in both groups varicella infection is severe. Passive immunization with hyperimmune immunoglobulin is an effective means of prevention when given within 72 hours of exposure. Every effort should be made to isolate patients with varicella and with herpes zoster. The management of pregnant women exposed to varicella is described in Table 11 (see Part 2 of this article). The problem with herpes simplex virus is more complex to manage (9). No therapy has yet become available to prevent the development of latency of HSV, subsequent recur- rences, and asymptomatic viral shedding. In addition, the absence of an effective means of immunization underlines the importance of alternative approaches which may help to prevent the transmission of the virus. Health-care providers consulted by patients with primary or recurrent genital herpes have an opportunity to reduce the spread of virus and to prevent at least some of its serious consequences. Patients should be informed about the clinical signs of the disease, the periodic nature of the infection, the potential for spreading the virus, and the ways of decreasing the risk of transmission to sex partners. Patients should be counselled to abstain 193 A WHO MEETING from intercourse from the first indication of the prodrome until the healing of the ulcerative lesion, and to use condoms during the asymptomatic intervening period in order to prevent transmission that might occur from virus shedding. This precaution might not be relevant in stable sexual relationships when both partners have evidence of previous HSV infection. In view of the close association of promiscuity and HSV-2 infections with cervical neoplasia, women with genital herpes should be advised to undergo yearly cyto- logical and colposcopical examinations of the cervix. Another major risk arising from genital HSV infection is neonatal infection, which is acquired by contact with infected genital tract secretions in the course of delivery. Intra- partum transmission of HSV from the mother to the infant may occur during primary or recurrent infections. As many as 30% of infants with neonatal HSV infection are born of mothers who are without symptoms at the time of delivery. Virological surveillance is therefore an important aspect in the antenatal care of females with a history of HSV infection or with sexual partners with a history of herpes simplex. It is generally agreed that weekly monitoring for the presence of virus in genital secretions and external genitalia should commence between the 32nd and 36th week of gestation. Women with evidence of asymptomatic virus shedding or clinical evidence of herpes lesions near term should be considered for Caesarian section, the only means of pre- vention of intrapartum transmission. The availability of a rapid, direct diagnostic test on clinical specimens would bring about a more rational approach to the problem. In situations where virological screening is not available, the management of pregnant women will have to be guided by the careful observation of prodromal signs and the appearance of herpes simplex lesions. In some environments a Pap test may be helpful. Infants born to infected mothers should be isolated in order to prevent the transmission of infection. LABORATORY DIAGNOSIS Recent advances in molecular biology have provided means for the rapid diagnosis of viral infections. The importance of this developing area has been recognized and the report of a WHO Scientific Group (41) is highly relevant to the diagnosis of herpesvirus infections. Methods for rapid diagnoses are continuing to improve and the technique of choice will change as details about the specificity and sensitivity of the tests become available. This report contains an assessment of the techniques now available. Laboratory diagnosis of herpesvirus infections can be made by virus isolation, by electron microscopy, or sometimes by light microscopic examination of clinical specimens. Diagnosis can also be made by demonstrating a significant rise in antibody titres to viral antigens in paired serum samples or by finding IgM antibodies to the virus in a single sample. The most commonly used methods are presented in Table 6. Virus isolation Virus isolation is, at present, generally more sensitive than the direct techniques commonly available for diagnosis and, when isolated from the lesion, provides evidence of its etiological role in the disease. Virus isolation is a good technique for diagnosing an asymptomatic infection as well as chronic and recurrent ones. The technique also permits precise identification of virus types or variants and the testing of antiviral drug sen- sitivity. However, there are several drawbacks to diagnosis by virus isolation. The technique is restricted to laboratories having cell culture facilities. The procedures are slow and the results are usually obtained within 2-3 days for HSV infections or after several days or even weeks in the case of CMV and VZV infections. The efficacy of isolation mainly 194 PREVENTION AND CON FROL OF HERPESVIRUS DISEASES Table 6. Methods commonly used for laboratory diagnosis of herpesvirus infections Applicability for: Methods HSV CMV VZV 1. Virus isolation t-" + + 2. Direct examination (a) Electron immune microscopy + + + (b) Cytology + + + (c) Immunofluorescence + - + (infected cells) (d) ELISA + - + (e) Counter-immune electrophoresis + 3. Serology (a) Complement fixation + + + (b) Neutralization + + + (c) Indirect immunofluorescence + + + (d) Indirect haemagglutination + + + (e) ELISA + + + (f) Radioimmunoassay + + + ' indicates most widely applied assays. Table 7. Preferred sources of specimens for virus isolation Viruses Sample CMV HSV VZV Skin (fluid from vesicles) _- ;38 ,t Throat swabs (saliva), bronchial aspiration or brushing + + +b Genital + + - Ocular - + - Urine + + + Breast milk + - - Blood cells or plasma + h + b + b 7 indicates the source of choice. b Immunocompromised patients. depends on the time of sampling after onset of the disease, specimens collected early in the course of illness yielding the highest isolation rates. Virus isolation rates are improved by transporting the sample under optimal conditions. Especially important is the presence of stabilizing substances such as protein in the transport medium, trans- porting the specimen at 4 °C, and reducing the transport time to a minimum. Virus isolation is most efficient in acute mucocutaneous infections since it may be difficult to obtain adequate specimens for severe deep-seated infections, e.g., in cases of encephalitis, hepatitis and disciform keratitis. Sources of clinical specimens which may yield viruses are listed in Table 7. The cell cultures of choice for isolating the herpesvirus are human diploid fibro- blasts (i.e., MRC-5). HSV-1 and 2 can also be readily isolated in continuous cell lines (such as Vero cells) or in rabbit kidney cells, but CMV and VZV do not replicate well in these cells. Typical cytopathogenic effects (CPE) develop in 1-4 days, 3-14 days, and 1-6 weeks in cultures inoculated with specimens containing HSV,' VZV and CMV, respectively. Further identification of the isolates can be done by immuno- logical tests such as neutralization, immunofluorescence, or ELISA. Poly- clonal antibodies have been used for identifying herpesviruses but it has been difficult to prepare such antibodies against CMV and VZV in animals. Thus, appro- priately standardized human sera have been used for identifying CMV. Recently, mono- clonal antibodies have been prepared to the herpesviruses and antibodies to HSV-1 and HSV-2 type-specific antigens (10) are now the reagents of choice for the identification and typing of isolates of these viruses. Monoclonal antibodies to CMV and VZV have been developed and should prove useful for identifying virus isolates. Diagnosis by direct examination of specimens Direct examination by electron microscopy of specimens collected from vesicle fluid (in the case of HSV and VZV) or urine (CMV) may reveal herpesvirus particles which provide evidence for a provisional diagnosis. Agglutination of particles with virus-specific anti- 195 A WHO MEETING bodies permits a specific diagnosis to be made. Cells infected with herpesvirus produce characteristic changes which include ballooning degeneration of the cells, ground-glass appearance with nuclei containing a basophilic inclusion and marginated chromatin, eosinophilic intranuclear inclusions, and multinucleated giant cells. Cells smeared on slides are fixed in methanol and stained with Giemsa or Wright stains. Cells stained by the Papanicolaou method also display changes when examined by light microscopy. Sections of biopsy specimens stained with haematoxylin and eosin can be examined for evidence of herpesvirus-induced change. When compared with virus isolation, diagnosis by electron microscopy or by light microscopy has been found to be less sensitive. A number of methods are available for direct detection of viral antigens in specimens (11). The antigen can be rapidly identified by direct or indirect ELISA. This system usually contains a catching and a detecting antibody. The detecting antibody may be labelled, or a labelled conjugate against the species of the detecting antibody may be used. This system is used for detection and typing of HSV and VZV in clinical specimens. The system has not yet been routinely used for the detection of CMV in clinical specimens, mainly because of the difficulties in obtaining hyperimmune sera against human CMV from animals. The fluorescent antibody (FA) technique is still the standard rapid viral diagnostic procedure for all viruses to which specific hyperimmune sera from animals can be obtained. It can be used also for the demonstration of HSV and VZV in clinical specimens and for the identifi- cation of all herpesviruses isolated in tissue culture. The advantages of the FA technique are that it allows localization of the virus in the infected cell and the results can be obtained rapidly. The disadvantages are that the technique is not yet applicable for the diagnosis of CMV infection because the infected cells in the urine are quickly destroyed. Highly specific antisera and good conjugates are required but are not always available. In addition, expen- sive equipment and technical experience are essential. Some specimens containing mucus and some skin lesions may give rise to unacceptable levels of nonspecific reaction. Solid phase radioimmunoassay (SPRIA) is similar to the ELISA with the exception that 1251 is used instead of enzyme label. This is a highly sensitive method but the reagents have a short shelf life and radioactivity presents a biological hazard. Antibody detection for herpesviruses Antibody detection is used for diagnosis of acute infection and of immunity status. With the majority of the conventional methods, mainly IgG antibodies can be reliably measured. These include complement fixation (CF), neutralization test (NT), the indirect haemag- glutination (IHA) tests, and the indirect immunofluorescence (IF) test. With these tests, for a specific serodiagnosis paired serum samples (taken 7-10 days apart) are required to document a significant antibody rise. In addition, the serodiagnosis of VZV or HSV infection may be obscured by a heterotypic antibody response. Since the IgM and IgA anti- bodies reflect recent antigenic stimulation, they have become of major importance for a specific rapid serodiagnosis with one serum sample. For IgM antibody detection, the IF technique has been used with questionable specificity, particularly in lower serum dilutions. With the indirect enzyme-linked immunoassay (ELISA) and also with radio- immunoassay, rapid techniques are available for measuring the IgM, IgA and IgG anti- bodies. The complement fixation test for antibody detection is also widely used and is nearly as sensitive as ELISA. However, its sensitivity depends on the potency of the CF antigens. The neutralization test has been the standard procedure. The addition of complement may enhance the seroactivity of the test. The major drawback of the NT is the need for tissue culture and the long observation period, especially in the case of CMV and VZV. 196 PREVENTION AND CONTROL OF HERPESVIRUS DISEASES 197 Difficulties in standardizing the neutralization procedure in order to obtain reproducible results is also recognized as a drawback and this topic was discussed at a joint meeting held by WHO and the Swedish Bacteriological Laboratory in Stockholm, 16-18 June 1982.b The fluorescent antibody technique is widely used and can detect different classes of antibodies for all herpesviruses. A particular test, the fluorescent antibody to membrane antigen (FAMA) test, has been specifically developed for the detection of antibody against VZV. The fluorescent conjugate reacts with the antibody, fixed by viral antigen expressed on the membranes of cells acutely infected with VZV. Using this method, the antibody is related to protection against exogenous VZV. Standardization of the FA test is difficult because of the reagents used, the presence of Fc receptor on the infected cells, and the sub- jectivity of the readings. In the indirect haemagglutination test the antibodies react with antigen coupled to red blood cells. IHA is a highly sensitive test but occasionally yields nonspecific results. The immune adherence haemagglutination assay (IAHA) has been developed for VZV antibody. This reaction depends on the activation of the C3 component of complement by the antigen-antibody complex. Indirect ELISA and RIA have been developed to detect classes of antibodies. ELISA is at present the most widely used test for demonstration of such antibodies. As with other tests, there are a number of pitfalls for the measurement of IgM antibodies. False positive results due to the presence of rheumatoid factors can be avoided by adsorption of sera with latex reagents or anti-IgG before testing or by the use of the mu-capture technique. With all IgM tests, there may be false low-positive results with unrelated antigens. ELISA tests for CMV, HSV and VZV are commercially available but their specificity and their sensitivity depend to a great extent on the quality of the antigen and on the specificity of the enzyme labels. At present, there is a great deal of variation among the commercially available kits. Related to the problem of assaying for type-specific antibodies to HSV-I and HSV-2 is the observation that the proteins of the viruses have both shared and unique antigenic regions. The possibility exists that each antigenic region may consist of a cluster of epitopes which vary in their relatedness to a similar region on the other virus serotypes. Studies with monoclonal antibodies capable of identifying individual epitopes have revealed intratypic variability in antigenicity of HSV-1 and HSV-2. In a study involving 36 virus isolates, over half of the monoclonal antibody preparations reacted anomalously (10). One anomalous pattern was the loss of reactivity to a type-specific or a type-common monoclonal antibody, while the second pattern was the reaction of a heterologous virus type with a type-specific monoclonal antibody. Where observed, 12-30Oo of variants lost such reactivity, while 11-16Vo cross-reacted. The implication of these observations is that the viruses contain mosaics of epitopes which are predominantly, but not exclusively, serotype-associated. The sharing of antigenic determinants by HSV-1 and HSV-2 complicates the use of standard antibody assays to distinguish easily and accurately between past infections with these two viruses. Sera obtained from patients infected only with HSV-1 or only HSV-2 usually have excess antibodies to the causative virus, as compared to the heterologous virus type. The picture is less clear for individuals infected with both types of virus. Recently, assays for type-specific antibodies have been described (12). The applicability of these assays in a diagnostic setting is now being carefully assessed. A special problem is the diagnosis of herpes simplex virus encephalitis. Antibody assays of serum and cerebrospinal fluid may be used to establish a diagnosis in the convalescent period by demonstrating a significant rise in antibody titre. However, brain biopsy is required early in the course of the disease when therapy is most beneficial (24). b Weekly epidemiological record, 57: 257, 261 (1982). 198 A WHO MEETING CHEMOTHERAPY Marketed drugs Idoxuridine (IDU), trifluridine or trifluorothymidine (TFT), and vidarabine (ara-A) (Fig. 1) are established drugs for the treatment of herpesvirus infections and have been licensed and marketed for several years (see Table 8) (13). Idoxuridine and trifluridine are only useful for topical treatment; they cannot be used systemically because of toxicity, especially for the bone marrow. These two drugs and vidarabine are widely used for the treatment of herpetic eye disease and idoxuridine has also been used as a 10% solution in dimethyl sulfoxide for the treatment of skin infections with herpes simplex and varicella- zoster virus. Vidarabine can be used systemically without prohibitive side-effects and has proved effective for the treatment of life-threatening infections, for example herpes ence- phalitis, herpes neonatorum, and varicella zoster in the immunocompromised host. More recently, another antiherpes drug, aciclovir (ACV) (Fig. 1), was licensed and marketed, and has been thoroughly reviewed (14). The earlier "first generation" anti- herpes drugs are non-selective inhibitors of DNA synthesis and their differential toxicity for viruses depends on the greater rate of DNA synthesis in virus-infected cells. By contrast, aciclovir is preferentially phosphorylated by the viral thymidine kinase (TK) to the monophosphate and subsequently converted to the triphosphate by cellular enzymes. The triphosphate of aciclovir then blocks DNA synthesis by inhibiting the DNA o O OHIN9 HN a tF3 HO HO H ~~~~~H lododeoxyuridine Trifluorothymidine(Idoxuridine, IOU) (Trifluridine, TFT O 0 HO-O- |J HO.O I OH Acycloguanosine lhydrypropoxymethyl- (Aclclovir. ACV) guanine (DHPG) HoH,+ ~~~~~~ l OH Bromovinyldeoxyuridine t BVOU NH2 N N) HOQ Carbocyclic analog of adenine arabinoside ( Cyclaradine) Br NH2 N 0 HO OH Fluoroiodoaracytosine ( FIAC) Cl C2H6 CH3-0O6 2)i 2H6 Arildone NH2 iN1 HO o OH Adenine arabinoside(Vidarabine,VraAA,A-A) 0 pC1,O-OO CH2 NH2 Glycyl ester of acyclo- guanosine(Glycyl-ACV) 0 Ht. CH3 HO OH Fluoromethylarauracil I FMAU) 0 O-P-Co 0* 3Nax Phosphonoformate(Foscarnet) Fig. 1. Structural formulae of antiherpes agents. Table 8. Licensed antiherpes drugs Drug Administration Idoxuridine (IDU) topical Trifluridine or trifluorothymidine (TFT) topical Vidarabine (Ara-A) topical and systemic (intravenous) Aciclovir (ACV) topical and systemic (intravenous and oral) dThd (dCyd) kinase / * ji HSV dTMP (MP vzv dTDP tv)DP dTT P X TP DNA polymerase --II T_ DNA ®DNA ®ACV, BVDU or FIAC Fig. 2. Mechanism of action of the nucleoside ana- logues, aciclovir (ACV), bromovinyldeoxyuridine (BVDU) and fluoroiodoaracytosine (FIAC) (see text). PREVENTION AND CONTROL OF HERPESVIRUS DISEASES 199 Table 9. Antiherpes drugs under investigation Drug Chemical name Foscarnet sodium trisodium phosphonoformate Glycyl-ACV aminoacyl esters of aciclovir DHPG" 9-(1,3-dihydroxy-2-propoxymethyl)guanine BVDU E-5-(2-bromovinyl)-2'-deoxyuridine FIAC 1-(2'-fluoro-2'-deoxy-13-D-arabinofuranosyl)-5-iodocytosine FMAU 1-(2'-fluoro-2'-deoxy-13-D-arabinofuranosyl)-5-methyluracil Cyclaradine carbocyclic 9-l&-D-arabinofuranosyladenine " DHPG is also known by other names, e.g., BW-759, BIOLF-62 and 2'-nor-2'-deoxyguanosine. polymerase and by acting as a DNA chain terminator. The sequence of events is illustrated in Fig. 2. A similar mode of action is shown by other recently developed antiherpes drugs such as bromovinyldeoxyuridine (BVDU) and fluoroiodoaracytosine (FIAC) (see below). Aciclovir has a very low toxicity and is excreted in the urine, having a half-life of 2.5 hours after intravenous administration. Renal toxicity occurred in early studies when the drug was given intravenously as a bolus, especially in poorly hydrated patients or those with renal disease. This problem can be solved by administering the drug by slow intra- venous infusion. Aciclovir has clearly shown a beneficial effect, compared with a placebo, in several conditions, such as: -topical treatment of herpetic keratitis; -topical/systemic (oral, intravenous) treatment of genital herpes, and, to a lesser extent, recurrent genital herpes; -systemic (intravenous) treatment of mucocutaneous herpes simplex virus infections in immunocompromised patients; -systemic (intravenous) treatment of varicella-zoster virus infections in immuno- compromised patients. As a rule, aciclovir is more effective the more serious the disease and the earlier it is given. Its effectiveness is most impressively demonstrated by the objective measurement of viral shedding. It has been shown to be effective when used systemically (orally, intra- venously) in the prophylaxis of HSV infections in immunosuppressed patients, i.e., bone- marrow transplant recipients. It may also be considered for the oral prophylaxis of recur- rent genital herpes in selected cases. New antiherpes drugs under development There are a number of other promising antiherpes drugs currently under development (see Table 9); their structures are shown in Fig. 1. The activity of antiviral drugs is usually measured in cell culture. Many variables affect this assessment and the most important are the virus strain, the multiplicity of infection (MOI), and the type of cells used as the substrate. Comparative tests for sensitivity are best performed in one laboratory using the same procedure (15, 16). The sensitivity to a drug in cell culture is only a rough guide to its usefulness in animals or man. A number of prodrugsc of aciclovir have been studied, e.g., glycyl-ACV. These have the ' A prodrug is a precursor of an active drug; the active drug is released from the prodrug in the presence of biological fluids or tissues. A WHO MEETING advantage of greater solubility in aqueous medium and also some of them are better absorbed orally. It is too early to know whether these drugs will prove superior to aciclovir, because toxicological and pharmacokinetic studies are necessary before large-scale clinical studies can be conducted. Another analogue of aciclovir is dihydroxypropoxymethylguanine (DHPG). This drug has a similar activity as aciclovir against HSV- I and HSV-2 in cell culture, but it is far more effective in animal models of herpes simplex. In cell culture DHPG is also more active than ACV against human cytomegalovirus and Epstein-Barr virus. BVDU is another interesting drug because of its highly potent and selective activity against HSV-1 and VZV. However, it is markedly less active against HSV-2. From phase I clinical trials BVDU appears to be a very promising drug for the topical treatment of herpetic keratitis and oral treatment of mucocutaneous HSV-1 and VZV infections in immunosuppressed patients, e.g., for treatment of varicella in leukaemic children and localized and disseminated zoster in cancer patients (17). According to phase I clinical trials, FIAC should be useful for the intravenous treatment of VZV infections in immuno- compromised patients (18). Double-blind placebo controlled clinical trials have shown foscarnet to be efficacious in the topical treatment of herpes labialis and genitalis. Unlike the nucleoside analogues (ACV, BVDU and FIAC), foscarnet does not require phosphorylation by the viral TK for activation (Fig. 2); it interacts directly with the viral DNA polymerase. Interferon. For a more detailed description of the therapeutic potentials of interferon, see the recent WHO report on this subject (19). Interferon has not yet achieved an established place in the treatment of herpesvirus infections. Human interferons (ca and ,B) have been tried in the treatment of herpetic keratitis, but they do not seem to offer any advantages compared with other recent anti- viral drugs. However, there may be benefits if interferon is used in combination with anti- viral drugs. Interferon has also been reported to be somewhat effective against varicella-zoster virus in immunosuppressed patients with malignant disease; despite the absence of comparative trials, interferon does not seem to be as effective as antiviral drugs. Studies on the prophylactic use of interferon in renal transplant patients have shown some promising results against HSV, CMV and EBV infections. Drug resistance The possible emergence of drug-resistant virus strains is a matter of concern for any form of chemotherapy. Thymidine kinase and DNA polymerase are the important enzymes at which resistance of HSV and VZV to aciclovir, BVDU, and the other nucleo- side analogues may develop (Fig. 2). Various HSV mutants have been isolated in the laboratory and they fall into four classes. There may be a total loss or a diminished level of expression of the TK; these are respectively the first and second classes of mutation. In general, these mutants have much reduced pathogenicity. The third class of mutation leads to a modified structure of the TK with markedly impaired ability to phosphorylate one compound, e.g., ACV, but with still tolerable competence to phosphorylate thymidine or other antiviral compounds. The fourth class of mutation occurring in the DNA polymerase gene leads to a lesser ability of the DNA polymerase to recognize the antiviral drug (as substrate or inhibitor) while still retaining its normal activity. Mutants of classes 3 and 4 give cause for concern because, if they were to emerge in practice, the resulting resistant virus might well be of full pathogenicity. At present, some clinical HSV isolates, especially from immunocompromised patients on ACV treatment have been found to be deficient in TK activity, but serious clinical resistance has not been 200 PREVENTION AND CONTROL OF HERPESVIRUS DISEASES 201 observed. However, the matter must be considered carefully, and surveillance of virus strains for sensitivity should be carried out. The study of these resistant strains is also important since it provides valuable information which should lead to more efficient chemotherapy. Limitations All the antiviral drugs discussed here achieve their antiviral effects by inhibition of viral DNA synthesis. They are active only in cells in which the virus is multiplying. Hence, none of these antiherpes agents will affect the virus during its latent state. In addition, the possible side-effects of the compounds during long-term administration and their potential hazards in pregnancy have not been assessed; until such information is available, the long-term administration of these compounds and their use in pregnant women should be approached with caution. (To be continued)