Viruses as Human Pathogen

 Chapter 8

Viruses as Human Pathogen

DNA Viruses

Viruses with Single-Stranded DNA Genomes

• The groups of viruses with single-stranded DNA genomes are contained in only one family, the parvoviruses, with only a single human pathogen type. The Geminiviridae, Circoviridae, and many other families have circular single-stranded DNA, but infect only plants and, more rarely, animals.

Parvoviruses

• This group’s only human pathogen, parvovirus B19, is the causative virus in erythema infectiosum (also known as “slapped cheek syndrome” or the “fifth disease”) in children and causes aplastic crisis in anemic patients. The virus also contributes to joint diseases, embryopathies, and tissue rejection following renal transplants. Diagnosis: serological (IgG and IgM) and PCR.

• Pathogen. The parvoviruses are among the smallest viruses with a diameter of 19–25 mm. They are icosahedral, nonenveloped, and their genome is in the form of single-stranded DNA (ssDNA). Some parvoviruses can only replicate in the presence of a helper virus (adenovirus or herpesvirus). Parvovirus B19, the only human pathogenic parvovirus identified to date, is capable of autonomic replication, i.e., it requires no helper virus. Some zoopathic strains also show this capability in rodents, dogs, and pigs.

• Pathogenesis and clinical picture. Parvovirus B19 replicates in the bone marrow in erythrocyte precursor cells, which are destroyed in the process. In patients already suffering from anemia (sickle-cell anemia, chronic hemolytic anemia), such infections result in so-called aplastic crises in which the lack of erythrocyte resupply leads to a critical shortage. In otherwise healthy persons, these infections usually run an asymptomatic course. They can, however, also cause a harmless epidemic infection in children, erythema infectiosum (“slapped-cheek syndrome” or “fifth disease”). This childhood disease, which used to be classified as atypical measles, is characterized by sudden onset of exanthem on the face and extremities. Certain forms of arthritis are considered complications of a parvovirus B19 infection. The virus also appears to cause spontaneous abortions in early pregnancy and fetal damage in late pregnancy (hydrops fetalis).

• Diagnosis. An enzyme immunoassay reveals antibodies of the IgG and IgM classes. During the viremic phase, at the onset of clinical symptoms, the virus can also be identified in the blood by means of electron microscopy or PCR. In-vitro culturing of the pathogen is not standard procedure.

• Epidemiology and prevention. The transmission route of human parvovirus B19 is not known. Droplet infection or the fecal-oral route, analogous to other parvoviruses, is suspected. Blood and blood products are infectious, so that multiple transfusion patients and drug addicts are high incidence groups. No specific prophylactic measures are recommended.

Viruses with Double-Stranded DNA Genome

• Viruses with double-stranded DNA genomes are classified in six families: papillomavirus, polyomavirus, adenovirus, herpesvirus, poxvirus, and hepadnaviral. Carcinogenic types have been found in all groups except the poxviruses (see Chapter 7, DNA tumor viruses).

Papillomaviruses

• The over 70 viral types in the genus Papillomavirus are all involved in the etiology of benign tumors such as warts and papilloma, as well as malignancies, the latter mainly in the genital area (cervical carcinoma). These organisms cannot be grown in cultures. Diagnosis therefore involves direct detection of the viral genome and histological analysis. Serology is less important in this group.

• anthogenesis and clinical picture. Papillomaviruses infect cells in the outer layers of the skin and mucosa and cause various types of warts by means of local cell proliferation. Specific virus types correlate with specific pathohistological wart types. Plantar and vulgar warts, flat juvenile warts, and juvenile laryngeal papilloma apparently always remain benign. By contrast, the genital warts caused by types 6 and 11 (condyloma a acuminata) can show carcinomatous changes. Of all papillomavirus-caused cervical dysplasia, 50% contain human papillomavirus (HPV) 16 and 20% HPV 18.

• All wart viruses induce primary proliferation of the affected cells with large numbers of viruses found in the cell nuclei. Whether a malignant degeneration will take place depends on the cell and virus type involved, but likely on the presence of cocarcinogens as well. In carcinomas, the viral DNA is found in integrated form within the host-cell genome, whereas in premalignant changes the viral genomes are found in the episomal state. Papillomaviruses possess oncogenes (E5, E6, and E7 genes) that bind the products of tumor suppressor genes: E6 binds the p53 gene product, E7 the Rb gene product.

• Diagnosis. Human papillomaviruses cannot be cultivated in vitro. They are detected and identified by means of histological analysis and, in malignancies in particular, by means of in-situ hybridization. Antibody assay results have a low significance level and these procedures are not standard routine.

• Epidemiology and prevention. Since viruses are produced and accumulate in wart tissues, papillomaviruses are transmissible by direct contact. Warts can also spread from one part of the body to another (autoinoculation). A certain level of prophylactic protection can be achieved with hygienic measures.

Polyomaviruses

• A medically important polyomavirus, the JC virus, causes progressive multifocal leukoencephalopathy (PML), a demyelinating disease that has become more frequent as a sequel to HIV infections, but is otherwise rare. The same applies to the BK virus, which affects bone marrow transplantation patients. Electron microscopy or PCR are the main diagnostic tools.

• Pathogens. The polyomaviruses can be divided into two groups: in one group are the SV40 and SV40-like viruses such as human pathogen JC and BK viruses. In the other are the true polyomaviruses such as the carcinogenic murine polyomavirus. The designations JC and BK are the initials of the first patients in whom these viraltypes were identified. There are also a number of other zoopathic oncogenic polyomaviruses. The name polyoma refers to the ability of this organism to produce tumors in many different organs.

• Pathogenesis and clinical picture. The JC and BK viruses are widespread: over 80% of the adult population show antibodies to them, despite which, clinical manifestations like PML are very rare. The viruses can be reactivated by a weakening of the immune defense system. The JC virus attacks the macroglia, especially in AIDS patients, to cause progressive multifocal PML, a demyelinating process in the brain with disseminated foci that is fatal within one year. The BK virus can cause hemorrhagic cystitis in bone marrow transplantation patients.

• Diagnosis. The JC and BK viruses can be grown in cultures, albeit with great difficulty and not for diagnostic purposes. Both can be detected with PCR and the BK virus can be seen under the electron microscope in urine. Antibody assays are practically useless due to the high level of generalized contamination.

• Epidemiology. Despite the high level of generalized contamination, the transmission routes used by the human polyomaviruses have not been clarified.

Adenoviruses

• There are a total of 41 types of adenoviruses, and they cause a wide variety of diseases. Influenza infections of the upper, less frequently the lower, respiratory tract and eye infections (follicular conjunctivitis, keratoconjunctivitis) are among the more significant clinical pictures. Intestinal infections are mainly caused by the only not culturable virus types 40 and 41. Diagnosis: antibody assay in respiratory adenovirus infections. Serology is not reliable in the eye and intestinal infections. It is possible to isolate the pathogens in cell cultures from eye infections. Enteral adenoviruses are detected in stool by means of electron microscopy, enzyme immunoassay, or passive agglutination.

• Pathogens. Adenoviruses are nonenveloped, 70–90 nm in size, and icosahedral. Their morphogenesis occurs in the cell nucleus, where they also aggregate to form large crystals. Their genome is a linear, 36–38 kbp double-stranded DNA. Adenoviruses got their name from the adenoidal tissues (tonsils) in which they were first identified.

• Pathogenesis and clinical picture. Adenoviruses cause a variety of diseases, which may occur singly or concurrently. The most important are infections of the upper (sometimes lower) respiratory tracts, the eyes, and the intestinal tract.

• Infections of the respiratory tract take the form of rhinitis or abacterial pharyngitis, depending on the virus type as well as presumably on the disposition of the patient. They may also develop into acute, influenzalike infections or even, especially in small children, into a potentially fatal pneumonia.

• The eye infections, which may occur alone but are often concurrent with pharyngitis, range from follicular conjunctivitis to a form of keratoconjunctivitis that may even cause permanent partial loss of eye.

• An important aspect of the intestinal infections is that the primary gastroenteritis forms are caused by the viral strains 40 and 41, which are difficult to culture.

• Diagnosis. Antibody assays in patient serum are the main approach taken in respiratory adenovirus infections. Serology is unreliable in the eye and intestinal infections, since hardly any antibodies are produced in response to such highly localized infections. It is possible to isolate the viruses that cause respiratory infections by inoculating cell cultures with pharyngeal material or bronchial secretion and with conjunctival smears in eye infections. Enteral adenoviruses, on the other hand, are hard to culture. The best approach to detecting them is therefore to subject stool specimens to electron microscopy, enzyme immunoassay, or passive agglutination methods.

• Epidemiology and prevention. Humans are the source of infection. Susceptibility is the rule. Generalized contamination of the population begins so early in childhood that adenovirus infections play a more significant role in children than in adults. Transmission of respiratory adenoviruses is primarily by droplet infection, but also as smear infections since the virus is also excreted in stool. Eye infections can be contracted from bathing water or, in the case of adenovirus type 8 in particular, iatrogenically from insufficiently sterilized ophthalmological instruments. The enteral infections are also transmitted by the fecal-oral route, mainly by contact rather than in water or food. Adenoviruses are the second most frequent diarrhea pathogen in children after rotaviruses.

Herpesviruses

• The viruses in this family all feature a practically identical morphology but show little uniformity when it comes to their biology and the clinical pictures resulting from infections. One thing shared by all herpesviruses is the ability to reactivate after a period of latency.

• The herpes simplex virus (HSV, two serotypes) is the pathogen that causes a vesicular exanthem (fever blisters, herpes labialis, or genitalis), encephalitis, and a generalized infection in newborns (herpes neonatorum).

• The varicella-zoster virus (VZV) causes the primary infection chickenpox, which can then recidivate as zoster (shingles).

• Cytomegalovirus (CMV) infections remain inapparent or harmless in the immunologically healthy, but can cause generalized, fatal infections in immunocompromised individuals.

• The Epstein-Barr virus (EBV) is the pathogen in infectious mononucleosis and is also implicated in lymphomas (including Burkitt lymphoma) and nasopharyngeal carcinomas.

• Human herpesvirus 6 (HHV 6) is the pathogen that causes three-day fever (exanthema subitum, roseola infantum).

• Diagnosis. Isolation, amplification culture, or direct detection can be used to diagnose herpes simplex, varicella-zoster, and cytomegaloviruses; antibody assays can be used for Epstein-Barr, human herpes 6 and 8, and varicella-zoster viruses; PCR can detect herpes simplex, varicella-zoster virus, cytomegalovirus, and human herpesvirus 6.

• Therapy. Effective and well-tolerated chemotherapeutics are available to treat herpes simplex, varicella-zoster virus, and cytomegalovirus (acyclovir, ganciclovir).

• Pathogen, pathogenesis, and clinical picture. The viral genome codes for about 90 proteins, categorized as “immediate early” (regulatory functions), “early” (DNA synthesis), and “late” (structural) proteins. Herpes simplex viruses are classified in types 1 and 2, which differ both serologically and biologically (host-cell spectrum, replication temperature). Initial infection with herpes simplex type 1 usually occurs in early childhood. The portal of entry is normally the oral mucosa (“oral type”) and the infection usually manifests as a gingivostomatitis. The viruses then wander along axons into the CNS, where they persist in a latent state in the trigeminal (Gasseri) ganglion. As with all herpesviruses, the pathogen remains in the macroorganism permanently after the primary infection. Following reactivation (endogenous recidivation), the viruses follow the same route back to the periphery, where they cause the familiar vesicular exanthem (“fever blisters,” herpes labialis. Despite established immunity, such recidivations can manifest repeatedly because the viruses wander within the nerve cells and do not enter.

• The initial infection with HSV type 2 normally affects the urogenital area (“genital type”) and can be contracted despite an existing HSV type 1 infection. HSV type 2 persists in the latent state in the lumbosacral ganglia or peripheral tissues, from where it causes episodes of manifest herpes genitalis. Neurological complications are very rare and more benign than in HSV type 1. On the other hand, infections of newborn children (herpes neonatorum), e.g., in cases of maternal genital herpes, are feared for their high lethality rate.

• Diagnosis. Cultivating the pathogen from pustule contents is the method of choice in labial and genital herpes. In an HSV encephalitis, the cerebrospinal fluid will contain few viruses or none at all. In such cases, they can only be cultivated from tissues (biopsy or autopsy material). Virus detection by means of cerebrospinal fluid PCR is worth a try.

• Epidemiology, prevention, and therapy. HSV type 1 is transmitted by contact, and possibly by smear infection as well. Contamination with HSV therefore begins in early childhood. Transmission of HSV type 2 usually occurs during sexual intercourse, so that infections are generally not observed until after puberty. No immune prophylaxis (vaccination) is currently available for HSV. Cyclobutanone is used prophylactically in immunosuppressed patients (see Chapter.

• Pathogen, pathogenesis, clinical picture. The VZ virus differs substantially from HSV, both serologically and in many biological traits. For instance, it can only be grown in primate cell cultures, in which it grows much more slowly and more cell-associated than is the case with HSV. No subtypes have been described.

• Pathogen, pathogenesis, clinical picture. CMV is characterized by a narrow spectrum of hosts, slow replication, frequently involving formation of giant cells and late, slow development of cytopathology. An initial infection with cytomegaly is inapparent in most persons, even in very early—perinatal or postnatal—infections. The virus apparently persists in the latent state in mononuclear cells. Reactivation can also run an asymptomatic course, but symptoms may also develop that are generally relatively mild, such a mononucleosislike clinical pictures, mild forms of hepatitis or other febrile illnesses. Droplet infection is the most frequent route of transmission, but smear infections and nursing infections are also possible. Generalized contamination with this pathogen (over 90% of the adult population is infected), frequent reactivation with, in some cases, months of continued excretion of viruses in saliva and urine and the wide variety of potential clinical pictures are all factors that often make it difficult to implicate CMVas the etiological cause of an observed illness. The virus infection can manifest as a sequel instead of a cause, for instance of a flulike illness. To labor the point somewhat, it could be said that the patient is not primarily ill due to a CMV infection, but rather has a florid CMV infection because he or she is ill.

• Diagnosis. Amplification cultures (p. 408f.) from saliva, urine, buffy coat, sue, or BAL (bronchoalveolar lavage) are a suitable method of confirming a florid CMV infection. In transplantation patients, the risk of a CMV manifestation can be estimated by immunocytochemical monitoring of the Compositive cell count in the peripheral blood (“antigenemia test”), since this count normally rises several days before clinical manifestations appear. Based on such an early warning, antiviral therapy can be started in time (ganciclovir, foscarnet). PCR results must be interpreted with a clear idea of how sensitive the method used can be, since the numbers of viruses found may be clinically insignificant. Hasty conclusions can result in “overdiagnosis,” above all in CMV-positive transplant recipients.

• Epidemiology, prevention, and therapy. CMV is transmitted by contact or smear infection, usually in childhood or adolescence. Immunosuppressed patients can be treated with hyperimmunoglobulin to provide passive immunity against infection or recidivation. Ganciclovir and foscarnet are therapeutically useful in transplantation, and particularly in AIDS patients, to combat CMV-induced pneumonia, encephalitis, and retinitis.

Epstein-Barr Virus (EBV)

• Pathogen pathogenesis, clinical picture. EBV infects only a narrow spectrum of hosts and replicates very slowly. It persists in a latent state in B lymphocytes and can lead to their immortalization and tumor transformation. EBV enters the body through the mucosa. It replicates in epithelial cells of the oropharynx or cervix and enters B lymphocytes, where it continues to replicate. This results in the clinical picture of infectious mononucleosis (kissing disease or Pfeiffer disease), which is characterized by fever and a generalized but mainly cervical swelling of the lymph nodes, typically accompanied by tonsillitis, pharyngitis, and some cases of mild hepatic involvement. This virus also persists in latency, probably for the life of the patient, in (immortalized) B cells.

• Chronic mononucleosis is characterized by antibodies to VCA and EA. The diagnostic procedures in lymphoproliferative diseases (see above) involve histology and cellular immunotyping.

• Epidemiology, prevention, and therapy. EBV is excreted in saliva and pharyngeal secretions and is transmitted by close contact (“kissing disease”). As with all herpesviruses the level of generalized contamination is high, with the process beginning in childhood and continuing throughout adolescence. Neither immunoprophylactic nor chemoprophylactic measures have been developed as yet. Lymphoproliferative diseases involving viral replication can be treated with acyclovir and ganciclovir.

Human Herpesvirus (HHV) 6

• Pathogen, pathogenesis, clinical picture. HHV-6 was isolated in 1986 in patients suffering from lymphoproliferative diseases and AIDS. The virus shows T-cell tropism and is biologically related to the cytomegalovirus. HHV-6 exists in two variants, HHV-6A and HHV-6B. The pathogenic implications of their reactivation have not yet been described.

• HHV-6B is the causal pathogen in exanthema subitem (roseola infantum), a disease that is nearly always harmless, characterized by sudden onset with high fever and manifests as a typical exanthem in small children. Reports of HHV-6-caused illness in adults are rare and the clinical pictures described resemble mononucleosis (EBV-negative mononucleosis). Apparently, however, this virus can also cause severe infections in bone marrow transplant patients (pulmonary and encephalitic infections). HHV-6A has not yet been convincingly implicated in any clinical disease.

Poxviruses

• The variola virus, which belongs to the genus Orthopoxvirus and is the causative agent in smallpox, was declared eradicated in 1980 after a WHO vaccination campaign. The vaccinia virus, used at the end of the 18th century by E. Jenner in England as a vaccine virus to protect against smallpox, is now used as a vector in molecular biology and as a hybrid virus with determinants from other viruses in experimental vaccines. Among the other orthopoxviruses found in animals, the monkeypox viruses are the main human pathogens. The animal pathogens parapoxviruses (milker’s nodules, orf) are occasionally transmitted to humans, in whom they cause harmless exanthems.

• Diagnosis. Orth poxviruses and parapoxviruses: electron microscopy. Molloscum contagiosum: histology.

• Pathogens. The viruses of the pox group are the largest viruses of all. At 230! 350 nm they are just within the resolution range of light microscopes. They have a complex structure (Fig. 8.7) and are the only DNA Viruses that replicate in a defined area within the host-cell cytoplasm, a so-called “virus factory”.

• The diseases smallpox (variola major) and the milder form alastrim (variola minor) now no longer occur in the human population thanks to a worldwide vaccination program during the 1970s. The last person infected by smallpox was registered in Somalia in 1977 and eradication of the disease was formally proclaimed in 1980. Since then, populations of the virus have been preserved in two special laboratories only.

• Pathogenesis and clinical picture. Variola viruses are transmitted aerogenic ally. The mucosa of the upper respiratory tract provides the portal of entry. From there, the pathogens enter the lymphoid organs and finally penetrate to the skin, where typical eruptions form and, unlike varicella pustules, all develop together through the same stages. The mucosae of the respiratory and intestinal tracts are also affected. Lethality rates in cases of smallpox (variola major) were as high as 40%. In alastrim (variola minor) the level was 2%, whereby the cause of death was often a bronchopneumonia.

• Infections with cowpox, orf, and milker’s nodule viruses are rare and usually harmless. The lesions remain localized on the skin (contact site), accompanied by a local lymphadenitis. These are typical occupational infections (farmers, veterinarians). The molluscum contagiosum virus is unusual in that in-vitro culturing of the virus has not succeeded to date. Infections with this virus do not confer immunity. The infection causes epidermal, benign tumors (“molluscum contagiosum warts”

• Diagnosis. The poxviruses group are relatively easy to recognize under an electron microscope in pustule contents, provided the pustules have not yet dried out or been superinfected with bacteria. Orth poxviruses and parapoxviruses can be differentiated morphologically, but the viruses within each genus share the same morphology. Molluscum contagiosum is diagnosed histologically.

• Epidemiology and prevention. Diseased humans were the sole viral reservoir of variola and alastrim. Transmission was direct and aerogenic and, although the virus is highly resistant even when desiccated, less frequent via contaminated objects (bed linens). The vaccinia virus does not occur in nature and any human infections are now accidental (laboratory infections). The zeotropic poxviruses are transmitted solely by means of contact with infected animals. Molluscum contagiosum is transmitted by interhuman contact.

Hantaviruses: Hepatitis B Virus and Hepatitis D Virus

• A hepatitis B virus (HBV) infection (see p. 385ff., replication) of the liver cells results in expression of viral antigen on the cell surface, followed by immunological cell damage with acute, possibly fulminant, chronic persistent or chronic aggressive hepatitis. The final stages can be liver cirrhosis or hepatocellular carcinoma. A concurrent or later superinfection by a defective, RNA-containing and HBV-dependent hepatitis D virus (HDV, delta agent) normally exacerbates the clinical course. Both viruses are transmitted in blood or body fluids, whereby even a tiny amount of blood may be enough to cause an infection.

• Diagnosis: immunological antigen or antibody assay in patient serum. The antigen or antibody patterns observed provide insights on the stage and course of the disease.

• Prevention: active immunization with HBV surface (HBs) antigen; concurrent postexposure passive immunization.

• Hepatitis B pathogen. The hepatitis B virus (HBV) is the main representative of the family of hantaviruses', Hepadnaviridae. The name of the family is an acronym of the disease caused by the virus and its genomic type. It causes a sometimes-chronic form of liver inflammation (hepatitis) and its genome consists of partially double-stranded DNA (hepadnaviral = hepatitis DNA virus). The replication cycle of the HBV includes a transient RNA phase (for details see Chapter 7, p. 385). The HBV possess an envelope made up f a cellular double lipid layer in which are integrated the hepatitis B surface (HBs) antigen, a 25 kDa polypeptide, and its precursor stages PreS1 (40 kDa) and PreS2 (33 kDa). (Fig. 8.10). This envelope encloses the actual capsid, which consists of the hepatitis B core (HBc) antigen with 21 kDa and contains the genome together with the DNA polymerase (a reverse transcriptase, p. 385). The complete, infectious virion, also known as a Dane particle after its discoverer, has a diameter of 42 nm, the inner structure 27 nm. The virus replicates in liver cells. The Dane particles and the HBs antigen, but not the HBc antigen, are released into the bloodstream, whereby the HBs antigen is present in two different forms, a filamentous particle approximately 22 ! 100 nm and a spherical form with a diameter of about 22 nm. A further viral protein is the HBe antigen, which represents a posttranslational, truncated form of the HBc antigen and is no longer capable of spontaneous capsid formation. It is also released from the hepatic cells into the blood.

• Hepatitis D pathogen. A certain percentage of HBV-infected persons, which varies geographically, are also infected by a second hepatitis virus discovered at the end of the seventies in Italy, the delta agent or hepatitis D virus (HDV). It was originally thought to be a new HBV antigen. In fact, it is an unclassified RNA virus that codes for the delta antigen. Its capsid consists of HBs antigen, i.e., HBV-coded material. For this reason, the virus can only replicate in persons infected with HBV (in this case the “helper virus”).

• Pathogenesis and clinical picture. The incubation period of hepatitis B is four to 12 weeks, followed by the acute infection phase, icteric, or anicteric course, once again with a variable duration of two to 12 weeks. The hepatic cell damage resulting from an HBV infection is not primarily due to cytopathic activity of the virus, but rather to a humoral and cellular immune response directed against the virus-induced membrane antigens (HBs, HBc) on the surface of the infected hepatocytes: 0.5–1% of those infected experience a fulminant, often lethal, hepatitis. In 80–90% of cases the infection runs a benign course with complete recovery and elimination of the HBV from the body. A chronic infection develops in 5–10% (see p. 393, persistent viral infections). Three forms are differentiated, but mixed forms are possible:

• healthy HBV carriers
• chronic persistent hepatitis (CPH) without viral replication, and finally
• chronic aggressive hepatitis (CAH) with viral replication and a progressive course

• Diagnosis. Hepatitis B is diagnosed by identifying the various HBV antigens or the antibodies directed against them. Both antigens and antibodies can be detected in patient blood using a solid phase test (enzyme immunoassay). The individual components manifest in specific patterns. Fig. 8.11 shows the sequence of phases in an uncomplicated hepatitis B infection, upon which the guiding principles in laboratory diagnosis of HBV infections are based.

• Epidemiology and prevention. Humans are the sole reservoir of HBV. Transmission is parenteral, either with blood or body fluids containing HBV (sexual intercourse) that come into contact with mucosa, lesions, or microlesions in the skin. In transmission by blood, the tiniest amounts contaminating syringe needles, ear-piercing needles, tattooing instruments, etc. suffice to produce an infection. Hepatitis B infections from blood transfusions have been greatly reduced by thorough screening of blood donors for HBs antigens, despite which patients receiving multiple transfusions or dialysis remain a highrisk group.

Picornaviruses

• Transmission of enteroviruses, parechoviruses, and hepatoviruses is by the fecal-oral route. The viruses first replicate in the intestine, from which location they reach their target organ with the bloodstream. Large numbers of inapparent infections are typical of this group. Rhinoviruses are transmitted by droplet infection and remain restricted to the upper respiratory mucosa.

• Diagnosis: enteroviruses and parechoviruses are diagnosed by isolation in cell cultures or with PCR, hepatitis A serologically (IgM) and rhinoviruses, if at all, by isolation.

• Prevention: basic polio immunization with dead or live vaccine; hepatitis A with dead vaccine; exposure prophylaxis with rhinoviruses.

• Pathogenesis and clinical pictures. The enteroviruses and parechoviruses are transmitted per os and replicate at first in the lymphoid tissue of the pharyngeal space, later mainly in the intestinal wall. They then reach their “target organs” via the bloodstream (e.g., CNS, muscles, heart, liver), followed by manifest organ infection, which, however, only develops in a small percentage of cases. Most infections run an asymptomatic course. Viremia is always present, so that even asymptomatic enterovirus and parechovirus infections confer effective immunity. The cases of manifest infection frequently run atypical courses with mild clinical symptoms. The same viral type can cause different symptoms and several different viral types can cause a given clinical symptom.

• Diagnosis. The available laboratory diagnostic tools include PCR or isolation of the virus from cerebrospinal fluid, pharyngeal smear, or lavage, with the best chances of success from stool. Serodiagnosis plays only a minor role.

• Epidemiology and prevention. Humans are the reservoir of the enteroviruses. Transmission is either direct (smear infection) or in food and water.

• Salk introduced a dead vaccine in 1954 for poliomyelitis prophylaxis (IPV, inactivated polio vaccine) consisting of three poliovirus types inactivated by formalin. Five years later, the live vaccine (OPV, oral polio vaccine according to Sabin) was introduced, which contains three live but no longer neurovirulent poliovirus strains, either singly or in combination. The WHO plan to eradicate poliomyelitis worldwide would seem feasible with this vaccine as demonstrated by its eradication in several countries including all of South Americ.

• Pathogen. The hepatitis A virus differs in some characteristics from enteroviruses, to which group it was long considered to belong. Growth in cell cultures requires long adaptation. Only one serotype is known to date.

• Pathogenesis and clinical picture. The clinical picture of hepatitis A, scaled epidemic or infectious hepatitis, differs in no major particulars from that of hepatitis B. The disease nearly always takes a benign course. Only a small number of fulminant (and sometimes lethal) or chronic courses have been described. The pathogenic process at first corresponds to that of the enteroviruses, whereby hepatitis A replicates in the intestine and then, after a brief viremic episode, attacks its target organ, the liver. Disease manifestation with this pathogen, unlike most of the enteroviruses but similar to hepatitis B, involves immunological processes.

• Diagnosis is based on IgM detection due to the early presence of these antibodies in patient serum, in fact so early that a lack of hepatitis A antibodies at the onset of clinical manifestations excludes hepatitis A.

• Epidemiology and prevention. Transmission is by food and water or in the form of smear infections. Infection with hepatitis A shows a clear north–south gradient: it has become virtually a travelers’ disease in central Europe. Imported cases frequently cause minor outbreaks in families or schools. Active immunization with an inactivated HAV vaccine is available.

Rhinoviruses

• Pathogens. The genomic organization and replication system of the rhinoviruses (117 serotypes found to date) generally match those of the enteroviruses, although they differ in that they are acid-sensitive and slightly denser.

• Pathogenicity and clinical picture. The rhinoviruses, the causative pathogens of the common cold, infect the mucosa of the nasopharyngeal space (nose and throat). They remain strictly localized there and do not cause generalized infections. In rare cases, mainly in children, they are known to cause bronchitis or bronchopneumonia as well. The clinical picture is often worsened by bacterial superinfection.

• Diagnosis. Laboratory diagnostics are only required in special cases of rhinovirus infection. The viruses can be grown in cell cultures.

• Epidemiology and prevention. Rhinoviruses are transmitted directly, for example by contaminated hands, and partly by droplet infection as well. Infective contacts between humans appear to involve mechanical inoculation (introduction into the nasopharyngeal space with fingers). Rhinoviruses occur worldwide, with pronounced proliferation in the winter months. The fact that everyone comes down with colds repeatedly is explained by the very brief immunity conferred by infection and the many different viral types involved. Experiments have shown that the infections are always exogenous, i.e., not reactivations due to cold, wetness, etc. The only conceivable prophylactic measure is to avoid large groups of people.

Astrovirus and Calicivirus; Hepatitis E

• Astroviruses, measuring 28–30 nm and caliciviruses, 30–35 nm, are enteritis pathogens in small children. Human pathogens in these groups include the Norwalk virus and hepatitis E virus (HEV). The latter occurs epidemically and endemically in Asian, Central American, and African countries. It is transmitted by the fecal-oral route, above all via drinking water, and causes relatively benign infections except in pregnant women. Hepatitis E is considered a traveler’s disease.

• Isolated cases and minor outbreaks of enteritis are typically attributed to unspecified viral infections. Besides unidentified bacterial infections, the viral pathogens that can cause such infections include adenovirus, rotavirus, astrovirus, and calicivirus, whereby the taxonomy of the latter two have not been confirmed.

Astroviruses

• Pathogen. The astrovirus is 28–30 nm in size and owes its name to its starlike appearance. It contains sense RNA with approximately 7 500 nucleotides and appears to have a replication strategy similar to that of the picornaviruses.

• Pathogenesis and clinical picture. Astroviruses that are animal and human pathogens are associated with episodes of diarrhea that nearly always run a harmless course. The etiological role of these viruses has still not been clarified. Astroviruses appear to possess only a low level of pathogenicity. It should be mentioned at this point that the role of viruses in enteritis is frequently exaggerated.

• Diagnosis. Detection by means of electron microscopy.

• Epidemiology. Astroviruses occur worldwide. They tend to infect young children and older persons weakened by other diseases.

Caliciviruses

• Pathogen. Caliciviruses are 30–35 nm, possess only one capsid protein and a polyadenylated, 7500-nucleotide RNA with a VPg at the 50 ends. The surface of the viruses has a characteristic structure with small, regular, calyxlike concavities that give the capsid the form of a Star of David. Caliciviruses are classified based on genomic similarities as either human caliciviruses (HuCV) or “small, round-structured viruses,” SRSV. This designation stems from their initial identification under the electron microscope as “small, round, virus particles.” The SRSVare grouped in two subtypes, I and II. Type I includes the Norwalk virus and a number of similar viruses named for their geographic venues, some with antigenicity differing from the Norwalk type.

• Clinical picture. Caliciviruses cause enteritis. Together with rotaviruses and adenoviruses, they are the most frequent viral enteritis pathogens in children, often causing minor epidemics during the winter months (“winter vomiting disease”)

• Diagnosis. Detection by means of electron microscopy or antigen assay in stool.

• Epidemiology. Two-thirds of the adult population in the temperate zone carry antibodies to the Norwalk virus. SRSVare regularly implicated in minor epidemics and family outbreaks. The transmission route of the Norwalk virus has been described: in addition to the fecal-oral route, water and uncooked foods are involved.

Hepatitis E Viruses

• Pathogen. An infectious inflammation of the liver endemic to Asia, Central America, and parts of Africa is apparently transmitted by the fecal-oral route. The RNA genome of the culprit agent has now been sequenced and the virus in question, the hepatitis E virus, has been classified with the caliciviruses. It occurs in at least 13 variants divided into three groups. In-vitro culturing of HEV has not succeeded to date.

• Pathogenesis and clinical picture. The clinical course of hepatitis E infections tends to be benign and resembles that of hepatitis A. It shows no chronicity. However, infections in the third trimester of pregnancy have a lethality rate of 10–40%.

• Diagnosis. The antibodies can be detected by means of an enzyme immunoassay. Apparently due to cross-reactions with other caliciviruses, the specificity of the results is uncertain. A diagnosis is often arrived at based on clinical evidence and medical history (travel to endemic areas).

• Epidemiology. HEV causes repeated outbreaks of considerable dimensions in the parts of the world mentioned above. The infections can be traced to contaminated drinking water. Hepatitis E is imported to central Europe as a traveler’s infection, although apparently less frequently than hepatitis A. No specific prophylactic measures exist.

Togaviruses

• Pathogen. The term togaviruses formerly included a variety of viruses, including what we now classify as the flaviviruses. As defined today, the togaviruses include the zoopathic pestiviruses, one species of rubivirus, the rubella virus and the alphaviruses with 25 species. The alphaviruses most important to travelers are the Chikungunya virus (Africa, Asia), the Sindbis virus (Africa, Asia, Australia), the Ross River virus (Australia, Oceania), and the Mayaro virus (South America), which are transmitted to humans by bloodsucking mosquitoes.

• Pathogenesis and clinical picture. The arthropodborne alphaviruses, zoonoses of the tropical and subtropical regions, frequently cause asymptomatic or benign infections with fever, exanthem, and joint pain. Occasionally, however, persistent arthralgia and polyarthritis (lasting months or even years) do occur, sometimes involving joint destruction. Even rarer, sequelae include encephalitis and meningoencephalitis with high lethality rates.

• Diagnosis. Serodiagnosis is the method of choice in suspected alphavirus and rubivirus infections. EIA methods are also available for IgM detection.

• Prevention. There are vaccines to protect against alphavirus infections and rubella. The main aim of rubella prophylaxis is to prevent rubella-caused embryopathies. Since 10–15% of young adults are still susceptible to rubella infections and a live vaccine with few side effects that confers reliable immunity is available, serial vaccination of children (boys and girls!) is done before puberty. The vaccine is tolerated so well that prior immune status checks are not required.

Flaviviruses

• Viruses in the flavivirus family (Flaviviridae) include the genera Flavivirus, Hepacivirus, and Pestivirus. Flaviviruses (the prototype being the yellow fever virus [Latin: flavus, yellow]) are transmitted by arthropods. They cause a biphasic infection that can have serious consequences (hemorrhagic fever with a high lethality rate). In southern and eastern countries, these viruses are significant human pathogens. Only one representative of this family, the tickborne encephalitis pathogen, is encountered in Europe. The hepaciviruses (hepatitis C [HCV] and hepatitis G viruses) are not arthropodborne. HCV is transmitted mainly in blood (transfusions, blood products, intravenous drug use) and is a frequent cause of chronic disease (70% of cases), including cirrhosis of the liver and hepatocellular carcinoma. The hepatitis G virus (HGV) is related to HCV and has not been characterized in detail as yet.

• Pathogen. The flavivirus family includes 63 species, among them the prototypic virus of the family, the yellow fever virus, and the pathogen that causes European tickborne encephalitis (spring-summer meningoencephalitis, SSME). lists the flaviviruses that cause significant travelers’ diseases.

• Pathogenesis and clinical picture. The arthropodborne flaviviruses cause diseases of different levels of severity. The infections are typically biphasic with an initial, not very characteristic phase including fever, headache, muscle pain, and in some cases exanthem (Dengue like disease). 

• Diagnosis. A flavivirus infection always involves viremia (transmission by bloodsucking arthropods!). The viruses can be isolated from blood by inoculating cell cultures or newborn mice. In autopsies of fatal cases they can be isolated from liver tissue. The viruses are labile by nature and identification can take time, for which reason the diagnostic focus is on serology (titer rise or IgM detection).

• Epidemiology and prevention. A cycle of infection involving a vertebrate host (mammals, birds) and a transmitting vector (bloodsucking mosquitoes and flies, ticks) has developed for most flavivirus infections. The cycles are efficient for the virus and relatively harmless for the host. The vertebrate host frequently shows few signs of disease and recovers from the infection after a brief viremia. During this period, the bloodsucking vector is infected, which thereafter remains a lifelong salivary secretor and thus infectious. In ticks, transovarian transmission of the virus is also possible. The human host is a dead end for the virus, not a normal component of the cycle. Exceptions to this are Dengue fever and urban yellow fever.

• Pathogen. A series of hepatitis infections following blood transfusions was observed that could not be identified as either hepatitis A or hepatitis B (p. 429) and were therefore designated as “non-A-non-B (NANB) hepatitis.” The discovery of the hepatitis C virus (HCV) by molecular biological means in 1988 was an elegant piece of work: RNA was extracted from the plasma of an infected chimpanzee, from which cDNA was produced using reverse transcriptase. The cDNA was then cloned, and the corresponding proteins expressed. About one million clones were tested for reactivity with sera from patients suffering from chronic NANB hepatitis. A protein was found by this method that reacted with antibodies to NANB, whereupon the corresponding cloned DNA was used as a probe to identify further overlapping gene segments. They belong to a flavivirus with approximately 10 kb sense RNA and several genotypes. A similar strategy led to identification of a further flavivirus that also causes hepatitis, now known as the hepatitis G virus (HGV).

• Pathogenesis and clinical picture. Hepatitis C resembles hepatitis B in many respects. One major difference is that it much more frequently produces a persistent infection (85%) and, in 70% of cases, develops into a chronic hepatitis, resulting in cirrhosis of the liver within 20 years and a hepatocellular carcinoma (HCC) in a further 10 years. The reason for the high level of viral persistence is thought to be a pronounced mutability facilitating evasion of the immune defenses (quasispecies of RNA viruses.

• Diagnosis of hepatitis C is done with antibody EIA using genetically engineered viral proteins. Western blot can be used to confirm the result. The RNA can be detected by means of RT-PCR and the course of therapy can be monitored with quantitative PCR.

Coronaviruses

• Infections with coronaviruses are widespread in humans and animals. Human pathogens include causative agents of rhinitis like infections and the virus of the “severe acute respiratory syndrome” (SARS), which first erupted in China in 2002. Diagnosis: serology or electron microscopy for common cold strains; PCR or isolation for SARS.

• Pathogen. The Coronaries family includes several viral species that can infect vertebrates such as dogs, cats, cattle, pigs, rodents, and poultry. The name (corona, as in wreath or crown) refers to the appearance of the viruses. One coronavirus species (human coronavirus, HuCV) is known since some time to be a human pathogen. It has at least two serotypes and probably a number of serological variants. In November 2002, a new coronavirus emerged in China and, after originally being mistaken as a new influenza recombinant, was identified as the causative agent of severe acute respiratory syndrome, or SARS, in spring 2003. Its origin, possibly from animals, is not known to date.

• Coronavirus Replication and Viral Maturation The coronavirus genome consists of the longest known, sense RNA strand exceeding 30 kb, which is integrated in the envelope in the form of a helical ribonucleoprotein. A hallmark of coronaviral RNA replication is the production of seven subgenomic mRNAs, each of which codes for one viral structural protein. The synthesis of progeny viral RNA takes place in association with specialized membrane structures, characterized as double-membrane vesicles. Viral maturation takes place in the rough endoplasmic reticulum after replacement of cellular proteins by viral proteins in the membranes. The viruses are then transported to the Golgi apparatus. The ensuing virus release mechanism is unknown. Recently, the receptor involved in the entry of the SARS virus into the cell was reported to be the angiotensin-converting enzyme 2 (ACE2).
• Pathogenesis and clinical picture. Common cold coronaviruses cause an everyday variety of respiratory infections, which are restricted to the ciliated epithelia of the nose and trachea. They are responsible for about 30% of common cold infections. The immunity conferred by infection, apparently IgA-dependent, is short lived. Reinfections are therefore frequent, whereby the antigenic variability of the virus may be a contributing factor. Various enteral coronaviruses with morphologies similar to the respiratory types have also been described in humans. Their pathogenicity, and hence their contribution to diarrhea, has not been clarified. The SARS virus is transmitted aerogenic Ally with an incubation time of two to 10 days. Clinically, fever and a marked shortness of breath is noted, developing into a severe atypical pneumonia with new pulmonary infiltrates on chest radiography. Shedding of virus is by respiratory discharges. Whether the virus present in other body fluids and excreta plays a decisive role for virus transmission is not yet clear.
• Diagnosis.The common-cold coronavirus can be grown in organ cultures of human tracheal tissue or in human diploid cells. Isolating the viruses for diagnostic purposes is not routine. Serodiagnosis (complement-binding reaction, immunofluorescence or enzyme immunoassay) and electron microscopy are feasible methods. The SARS virus can be identified by PCR or isolated in the Vero cell line.
• Epidemiology and prevention. In November 2002, an outbreak of atypical pneumonia, later termed SARS, occurred in the southern Chinese city of Guangzhou (Guangdong Province). Only in February of 2003, the world was alerted about the lung disease, shortly before it escaped China, when a Guangdong resident in a Hong Kong hotel transmitted it to other guests who spread it to Toronto, Hanoi, Singapore, and elsewhere. Transmission of the virus is by droplets, but close contact (“household transmission”). 

Retroviruses

• Diagnosis: HIV infections are routinely detected by serology (antibodies or viral antigen). The circulating virus count (viral load) is determined by means of quantitative RT-PCR. The AIDS diagnosis is a clinical procedure that presupposes positive confirmation of HIV infection.

• Prevention: exposure prophylaxis when contact with blood is involved (drug addicts, healthcare staff) and sexual intercourse. Postexposure prophylaxis and prophylaxis in pregnancy with chemotherapeutics.

• Pathogen. The Retrovir family is the classification group for all RNA viruses with reverse transcription of RNA to DNA in their reproductive cycles (RNA-dependent DNA synthesis) (p. 385). Only zoopathic retroviruses were known for many years. These viruses cause various kinds of tumors in animals. In 1980, retroviruses were also discovered in humans. This virus family includes seven genera, three of which play significant roles in human medicine:

• HTLV-BLV retroviruses, including HTL virus types I and 
• II and the bovine leukemia virus. Sumair's
  
  
  
  
  , which only occur in animals, two of which are (probably) from humans.
• Lentiviruses, with the human pathogens HIV 1 and 2, maedivirus (pneumonia), and visnavirus (encephalomyelitis) in sheep, viruses affecting goats and horses, and animal immune deficiency viruses

Subviral Pathogens: Viroids and Prions

• Viroids are Phyto pathologically significant, noncoding RNA molecules that interfere with cellular regulation as antisense RNA. The hepatitis D virus has some structural similarity to viroids.

• Prions consist of a cell-coded protein (PrP: prion protein) altered in its conformation and by point mutations. They are infectious and can cause normal cellular PrP to assume the pathological configuration. They cause the spongiform encephalopathies (Creutzfeldt-Jakob disease, CJD) in the classic and new variants (nvCJD), the Gerstmann-Stra¨ussler-Scheinker (GSS) syndrome, and animal diseases (scrapie, BSE) characterized by neuronal vacuolization and loss and so-called amyloid plaques.

Viroids

• iroids were discovered at the end of the sixties during investigations of plant diseases. They consist of infectious, naked ssRNA in closed circular form with extensive base-pairing to form a rod-shaped strand 50 nm long. This RNA is 10 times smaller than the smallest viral nucleic acid and comprises, depending on the type, only 250–350 nucleotides. It does not function as mRNA and does not code for proteins. Its mode of replication is unknown, but certainly involves cellular enzymes.

• Viroids cause a number of plant diseases with considerable economic impact. The following hypothesis is currently under discussion to explain their pathogenicity mechanism: viroids possess complementary sequences to cellular 7S RNA, comprising, together with six proteins, the “signal recognition particle.” This particle controls the posttranslational membrane insertion of proteins. Viroids can thus interfere as “antisense (or interfering) RNA” with the function of 7S RNA and thus with membrane formation.

Prions

• Pathogen. Attention was first drawn to certain encephalopathic agents whose physical properties differed greatly from those of viruses. For instance, they showed very high levels of resistance to sterilization and irradiation procedures. It was later determined that these pathogens—in complete contrast to viruses and viroid require only protein, and no nucleic acid, as the basis of their infectivity and pathogenicity. This gave rise to the term “prion” for “proteinaceous infectious particle.” An intensive search for nucleic acid in the “particles” was fruitless.

• Prions are misfolded forms of a cellular protein. They consist of only a single protein (PrP, prion protein), which naturally occurs, for example, on the surface of neurons. The region coding for this protein of approx. 35 kDa is located in a single exon and is derived from a cellular gene expressed in both healthy and diseased brains. Disease-associated PrP (the best-known prion is the scrapie pathogen, the protein of which is called PrPsc [sc for scrapie]), is a mutant, slightly shortened (27–30 kDa) form of the normal PrPc (c for cell). It differs from normal PrPc in its altered configuration, its nearly complete resistance to proteases and in the fact that it tends to accumulate inside the cell.

• Pathogenesis. Infectious PrPsc can transform naturally occurring PrPc into PrPsc, resulting in an autocatalytic chain reaction in which mainly the pathological protein is produced. This is why mice lacking the gene for PrP (genetically engineered “knockout mice”) cannot be infected with the pathological Prisca prion. Deposits of large amounts of the pathological protein in the form of so-called amyloid plaques are visible under a microscope in brain tissue from infected humans and animals.

• Diagnosis. The diagnostic procedure is histological. Since there is no immune response to the pathological PrP, Sero diagnostic methods are useless. The pathological protein can, however, be detected in lymphoid tissue biopsies using monoclonal antibodies.

• Epidemiology. CJD, which occurs sporadically (one case per million inhabitants per year) is produced anew in every case by mutations in Props. The disease can be transmitted iatrogenically (brain electrodes, corneal transplants). The pathogenicity of the GSS prion (PrPgss) is based on a single amino acid change. Genetic factors also appear to have a predisposing effect in view of the existence of familial forms of GSS. Kuru is a disease that was spread in New Guinea by cannibalistic rites, probably originating with a case of CJD. Kuru no longer occurs today.