Viruses

1. H E R P E S V I R U S E S ; H U M A N P A P I L L O M A V I R U S
DNA VIRUSES

3. HSV type 1 (HSV-1) and type 2 (HSV-2) are distinguished by two main criteria:
antigenicity and location of lesions. Lesions caused by HSV-1 are, in general,
above the waist, whereas those caused by HSV-2 are below the waist.
Diseases
HSV-1 causes acute gingivostomatitis, recurrent herpes labialis (cold sores),
keratoconjunctivitis (keratitis), and encephalitis, primarily in adults. HSV-2
causes herpes genitalis (genital herpes), neonatal encephalitis and other
forms of neonatal herpes, and aseptic meningitis. Infection by HSV-1 or HSV-2
is a common cause of erythema multiforme.
Important Properties
HSV-1 and HSV-2 are structurally and morphologically indistinguishable. They
can, however, be differentiated by the restriction endonuclease patterns of
their genome DNA and by type-specific monoclonal antisera against
glycoprotein G. Humans are the natural hosts of both HSV-1 and HSV-2.
HERPES SIMPLEX VIRUSES (HSV)

4. Replicative Cycle
The cycle begins when HSV-1 binds first to heparan sulfate on the cell surface
and then to a second receptor, nectin.
Following fusion of the viral envelope with the cell membrane, the
nucleocapsid and the tegument proteins are released into the cytoplasm.
The viral nucleocapsid is transported to the nucleus, where it docks to a
nuclear pore and the genome DNA enters the nucleus along with tegument
protein VP16. The linear genome DNA now becomes circular.
VP16 interacts with cellular transcription factors to activate transcription of
viral immediate early (IE) genes by host cell RNA polymerase. IE mRNA is
translated into IE proteins that regulate the synthesis of early proteins such as
the DNA polymerase that replicates the genome and thymidine kinase.
These two proteins are important because they are involved in the action of
acyclovir, which is the most important drug effective against HSV.
Note that early protein synthesis by HSV can be subdivided into two
categories: immediate early and early. Immediate early proteins are those
whose mRNA synthesis is activated by a protein brought in by the
incoming parental virion (i.e., no new viral protein synthesis is required for
the production of the five immediate early proteins). The early proteins, on
the other hand, do require the synthesis of new viral regulatory proteins to
activate the transcription of their mRNAs.

5. The viral DNA polymerase replicates the genome DNA, at which time
early protein synthesis is shut off and late protein synthesis begins. These
late, structural proteins are transported to the nucleus, where virion
assembly occurs.
The virion obtains its envelope by budding through the nuclear
membrane and exits the cell via tubules or vacuoles that
communicate with the exterior.
In latently infected cells, such as HSV-infected neurons, circular HSV
DNA resides in the nucleus and is not integrated into cellular DNA.
Transcription of HSV DNA is limited to a few latency-associated
transcripts (LATS). These noncoding, regulatory RNAs suppress viral
replication. Reactivation of viral replication can occur at a later time
when the genes encoding LATS are excised.

6. Laboratory Diagnosis
An important diagnostic procedure is isolation of the virus from the lesion by
growth in cell culture. The typical cytopathic effect occurs in 1 to 3 days,
after which the virus is identified by fluorescent antibody staining of the
infected cells or by detecting virus-specific glycoproteins in enzymelinked
immunosorbent assays (ELISAs). HSV-1 can be distinguished from HSV-2 by
using monoclonal antibody against glycoprotein G often in an ELISA test.
A rapid presumptive diagnosis can be made from skin lesions by using the
Tzanck smear, in which cells from the base of the vesicle are stained with
Giemsa stain. The presence of multinucleated giant cells suggests herpesvirus
Infection
If herpes encephalitis is suspected, a rapid diagnosis can be made by
detecting HSV DNA in the spinal fluid by using a polymerase chain reaction
(PCR) assay. The PCR assay is more sensitive than viral culture. The diagnosis
of neonatal herpes infection typically involves the use of viral cultures
or PCR assay.
Serologic tests such as the neutralization test can be used in the diagnosis of
primary infections because a significant rise in antibody titer is readily
observed. However, they are of no use in the diagnosis of recurrent infections
because many adults already have circulating antibodies, and recurrences
rarely cause a rise in antibody titer.

7. Diseases
Human papillomavirus (HPV) causes papillomas, which are benign tumors of
squamous cells (e.g., warts on the skin). Some HPV types, especially types 16
and 18, cause carcinoma of the cervix, penis, and anus.
Important Properties
Papillomaviruses are nonenveloped viruses with doublestranded circular DNA
and an icosahedral nucleocapsid. The HPV genome has seven early genes (E1–
E7) and two late genes (L1 and L2). The early genes encode proteins
involved in the synthesis of viral mRNA and in the replication of the progeny
DNA genomes, and the late genes encode the structural proteins of the
progeny virions.
Two of the early genes, E6 and E7, are implicated in carcinogenesis. They
encode proteins that inactivate proteins encoded by tumor suppressor genes in
human cells (e.g., the p53 gene and the retinoblastoma [RB] gene,
respectively). Inactivation of the p53 and RB proteins is an important step in the
process by which a normal cell becomes a cancer cell.
There are at least 100 types of papillomaviruses, classified primarily on the basis
of DNA restriction fragment analysis. There is a pronounced predilection of
certain types to infect certain tissues. For example, skin warts are caused
primarily by HPV-1 through HPV-4, whereas genital warts (condylomata
acuminata) are usually caused by HPV-6 and HPV-11. Approximately 30 types
of HPV infect the genital tract.
PAPILLOMAVIRUSES

8. Replicative Cycle
After attachment and uncoating, the genome DNA moves to the nucleus.
Messenger RNA is synthesized by host cell RNA polymerase with early viral
protein E2 acting as a transcriptional activator. Early viral protein E1 acts as a
helicase that separates the DNA strands of the incoming viral genome. This
allows the host cell DNA polymerase to synthesize the progeny DNA
genomes. The initial progeny genomes are maintained as episomes in the
nucleus. Most of the synthesis of progeny viral DNA occurs in conjunction
with cellular DNA synthesis during S phase.
Late mRNA’s encode both the major structural protein (L1) and the minor
structural protein (L2). L1 protein comprises the capsid of HPV virions. L1 has
the ability to selfassemble into capsids in vitro and it is this form that is the
immunogen in the HPV vaccine. L2 protein aids in the packaging of genome
DNA into the progeny virions as well as in uncoating the genome when they
infect the next cell.
In human tissue, infectious virus particles are found in the terminally
differentiated squamous cells rather than in the basal cells (Figure 38–1A).
Note that HPV initially infects the cells of the basal layer in the skin, but no
virus is produced by the basal cells. Rather, infectious virions are produced
by squamous cells on the surface, which enhances the likelihood that
efficient transmission will occur.
In malignant cells, viral DNA is integrated into host cell DNA in the vicinity of
cellular proto-oncogenes, and E6 and E7 are overexpressed.

9. Laboratory Diagnosis
Infections are usually diagnosed clinically. The presence of
koilocytes in the lesions indicates HPV infection. A polymerase
chain reaction (PCR)–based test can be used to detect the
presence of the DNA of 14 high-risk genotypes, including HPV-16
and HPV-18. Pap Smears are done to detect cervical cancer.
Papanicolaou stain
Koilocytes, also known as halo cells, are a type of epithelial cell
that develops following a human papillomavirus (HPV)
infection. Koilocytes are structurally different from other
epithelial cells. For instance, their nuclei, which contain the cell's
DNA, are an irregular size, shape, or color.

10. RNA VIRUSES
PICORNAVIRUSES (POLIOVIRUS );RETROVIRUSES (HIV)

11. The picornavirus family includes two groups of medical importance:
the enteroviruses and the rhinoviruses. Among the major enteroviruses
are poliovirus, Coxsackie viruses, echoviruses, and hepatitis A virus.
Enteroviruses infect primarily the enteric tract, whereas rhinoviruses are
found in the nose and throat (rhino = nose).
Picornaviruses are small (20–30 nm) nonenveloped viruses composed
of an icosahedral nucleocapsid and a singlestranded RNA genome.
The genome RNA has positive polarity (i.e., on entering the cell, it
functions as the viral mRNA). There is no polymerase within the virion.
Picornaviruses replicate in the cytoplasm of cells. They are not
inactivated by lipid solvents, such as ether, because they do not have
an envelope.

12. POLIOVIRUS
Disease
This virus causes poliomyelitis.
Important Properties
The host range is limited to primates (i.e., humans and nonhuman
primates such as apes and monkeys). This limitation is due to the
binding of the viral capsid protein to a receptor found only on
primate cell membranes. However, note that purified viral RNA
(without the capsid protein) can enter and replicate in many
nonprimate cells—the RNA can bypass the cell membrane receptor
(i.e., it is ―infectious RNA‖).
There are three serologic (antigenic) types based on different
antigenic determinants on the outer capsid proteins. Because there
is little cross-reaction, protection from disease requires the presence
of antibody against each of the three types.

13. Replicative Cycle
The virion interacts with specific cell receptors on the cell membrane
and then enters the cell. The capsid proteins are then removed. After
uncoating, the genome RNA functions as mRNA and is translated into
one very large polypeptide called noncapsid viral protein 00. This
polypeptide is cleaved by a virus-encoded protease in multiple steps to
form both the capsid proteins of the progeny virions and several
noncapsid proteins, including the RNA polymerase that synthesizes the
progeny RNA genomes. Replication of the genome occurs by synthesis
of a complementary negative strand, which then serves as the
template for the positive strands. Some of these positive strands function
as mRNA to make more viral proteins, and the remainder become
progeny virion genome RNA. Assembly of the progeny virions occurs by
coating of the genome RNA with capsid proteins. Virions accumulate in
the cell cytoplasm and are released upon death of the cell. They do
not bud from the cell membrane.

14. Laboratory Diagnosis
The diagnosis is made either by isolation of the virus or by a rise in
antibody titer. Virus can be recovered from the throat, stool, or spinal
fluid by inoculation of cell cultures.
The virus causes a cytopathic effect (CPE) and can be identified
by neutralization of the CPE with specific antisera.

15. HUMAN
IMMUNODEFICIENCY VIRUS
Disease
Human immunodeficiency virus (HIV) is the cause of acquired
immunodeficiency syndrome (AIDS). Both HIV-1 and HIV-2 cause
AIDS, but HIV-1 is found worldwide, whereas HIV-2 is found primarily
in West Africa.
Important Properties
HIV is one of the two important human T-cell lymphotropic
retroviruses (human T-cell leukemia virus is the other). HIV
preferentially infects and kills helper (CD4) T lymphocytes, resulting
in the loss of cell-mediated immunity and a high probability that the
host will develop opportunistic infections. Other cells (e.g.,
macrophages and monocytes) that have CD4 proteins on their
surfaces can be infected also.

16. The genome of HIV consists of two identical molecules of single-
stranded, positive-polarity RNA and is said to be diploid. The HIV
genome is the most complex of the known retroviruses. In addition to
the three typical retroviral genes gag, pol, and env, which encode
the structural proteins, the genome RNA has six regulatory genes. Two
of these regulatory genes, tat and rev, are required for replication,
and the other four, nef, vif, vpr, and vpu, are not required for
replication and are termed ―accessory‖ genes. The gag gene
encodes the internal ―core‖ proteins, the most important of which is
the p24 protein. It is important medically as it is the antigen in the
initial serological test that determines whether the patient has
antibody to HIV (i.e., has been infected with HIV).

19. Laboratory Diagnosis
The presumptive diagnosis of HIV infection is often made by the
detection of antibodies in the patient’s serum to the p24 protein of HIV
using the enzyme-linked immunosorbent assay (ELISA) test. Because
there are some falsepositive results with this test, the definitive diagnosis
is made by Western blot (also known as Immunoblot) analysis, in which
the viral proteins are displayed by acrylamide gel electrophoresis,
transferred to nitrocellulose paper (the blot), and reacted with the
patient’s serum. If antibodies are present in the patient’s serum, they
will bind to the viral proteins (predominantly to the gp41 or p24.
protein). Enzymatically labeled antibody to human IgG is then added.
A color reaction reveals the presence of the HIV antibody in the
infected patient’s serum.
OraQuick is a rapid, screening immunoassay for HIV antibody that uses
an oral swab sample in an ELISA-type test that can be done at home.
Results are available in 20 minutes. Positive results for HIV antibody
require confirmation by a Western blot test.

20. The PCR test is a very sensitive and specific technique that can be used
to detect HIV DNA within infected cells. The presence of HIV can be
detected during acute infection by the plasma HIV RNA assay (viral
load), as viremia is typically high at this early stage. Also useful for the
diagnosis of early infections is the HIV antigen/ antibody ―Combo‖ test
that detects the presence of p24 antigen as well as antibodies to both
HIV-1 and HIV-2.
This combination test is useful for the diagnosis of early infections
because p24 antigen is typically detectable earlier in infection than
antibody.