1.            What is the structure of papilloma viruses? Genome type? Enveloped? Symmetry?

 HPV is a non-enveloped, icosahedral virus with a circular, double-strand DNA genome.

Distinguish permissive vs non-permissive cells as they relate to HPV.

 Permissive cells allow for lytic replication of the virus.  Non-permissive cells do not produce progeny virus (late proteins are not expressed), but instead are transformed by the E6 and E7 early proteins.  Benign tumors contain many copies of the virus genome as extra-chromosomal circular copies.  Malignant tumors contain integrated portions of the virus genome which results in higher expression of the E6 and E7 proteins.

What role do early (“E”) proteins play in virus replication? In transformation?

 The early proteins promote viral gene transcription and virus genome replication.  E6 binds p53 (tumor suppressor protein) and E7 binds Rb (another tumor suppressor protein), disrupting the normal cell cycle leading to immortalization of the cells.

 Describe the morphogenesis of a wart.  Why are HPV infections difficult to eliminate?

 HPV infects the germinal cells in the basal layer of the skin.  These cells are non-permissive for HPV, but serve as a permanent reservoir of HPV genomes.  This is why HPV is difficult to cure and warts often return.  As the germinal cells differentiate into keratinocytes, they become permissive for HPV.  As a result, when a wart is formed on the skin’s surface, it is producing virus and is infectious.  This can lead to multiple warts in a localized area.

Common and plantar warts: verruca vulgaris

      Flat warts: verruca plana

      Anogenital warts: condyloma acuminate and condyloma plana

Which HPV genotypes are considered low risk for cancer? High risk?

Low risk: HPV 6 and 11

      High risk: HPV 16, 18, and 31

 How is SPI detected in women? What are koilocytotic cells?  Why are cervical cancer deaths lower in the US compared to developing nations?

 SPI is suspected in abnormal Pap smears, and affected cells can be detected by brushing 5% acetic acid on mucosal surfaces and looking for white patches.  Koilocytotic cells can be seen in abnormal Pap smears and appear to have a “vacuolated” or washed out cytoplasm surrounding the nucleus.  Cervical cancer is the number 2 cause of cancer deaths in women world-wide, but the incidence is much lower in developed nations because of Pap screening, and hence, early detection.  HPV vaccination will further reduce the incidence of cervical cancer.

 What are important co-factors (in addition to HPV infection) for the development of cervical cancer?

 Co-carcinogens for cervical cancer include herpes simplex virus genital infection and cigarette smoking.

 What comprises Gardasil, the HPV vaccine?

 Gardasil is a tetra-valent vaccine composed of the L1 (late 1) capsid protein from HPV 6, 11, 16, and 18.  The L1 proteins are expressed in yeast where they form capsid-like structures by self-assembly and are purified to serve as antigens in the vaccine.  The vaccine has nearly 100% efficacy against HPV 6, 11, 16, and 18, and may protect somewhat from closely related strains.  A di-valent vaccine that contains yeast-expressed L1 proteins from HPV 16 and 18 is approved for use in Europe.

 Describe treatment methods for HPV infections

 Easily visible warts will usually go away on their own after several months or a couple of years.  Common warts are typically destroyed by cryosurgery or the use of denaturing agents such as bi- or trichloroacetic acid.  Topical agents such as podofilox (anti-mitotic agen) or imiquimod (stimulates a localized innate immune response) can be used for anogenital warts.  Intralesional interferon has also been approved for treatment of common warts.  Plantar warts may need surgical excision due to their location on weight-bearing areas of the feet (painful to stand).  Loop electrosurgical excision procedures (LEEP) of the cervix may be necessary in cases of abnormal Pap smears if the abnormality is graded sufficiently high on a dysplasia index. 

How do herpesviruses enter cells?

Herpesviruses fuse directly with the plasma membrane of cells in a pH-independent process.

In a Tzanck smear, multinucleated, giant cells (syncitia) containing nuclear inclusion bodies (the result of virus production in the nucleus) are observed microscopically in scrapings taken from a vesicle.  Syncitia are formed because the same viral envelope glycoproteins that mediate virus fusion and entry are present on the surface of infected cells late in infection.  The glycoproteins are able to fuse the infected cell plasma membrane with neighboring cell membranes, leading to giant cells with more than one nucleus.

 HSV 1 is principally associated with which diseases? HSV 2?

HSV 1: gingivostomatitis, fever blister, keratitis and other eye infections, encephalitis

      HSV 2: genital lesions, meningitis, neonatal infections (skin, eyes, mouth, disseminated, CNS including encephalitis)

 Where do herpes simplex 1 or 2 reside during latency?  What is the role of LAT?  What are factors that lead to virus reactivation?

 Latency is established in peripheral sensory neurons, primarily of the trigeminal and sacral ganglia.  The genomes are intact and are found extrachromosomally (they do not integrate into our chromosomes).  LAT is the latency-associated transcript; it does not encode a protein.  Instead, it is believed to act as a silencing RNA to block apoptosis of the neuron.  This is important because virus infection would normally induce apoptosis, but it is imperative (to the virus) that the neuron survive to serve as a life-long reservoir of the virus.  Reactivation occurs at times of stress (sunburn, illness, exams, etc) when cell-mediated immunity drops.  The lesion, for example a cold sore, remains small because neutralizing antibody is maintained at fairly high titers (especially during periods of frequent reactivation).  Lesions recur at the same site because these are the areas innervated by the latently-infected neurons

Compare and contrast herpes genital lesions from those of syphilis and chancroid.

 Herpes                 Syphilis               Chancroi

Multiple, smaller vesicles     One papule             Multiple pustules

   Superficial, no induration    Sharply demarcated   Deep, undefined ed

   Tender, red edges               Painless, purulent baseTender, purulent base

Other than skin lesions, what other symptoms are associated with herpes genital infections?

 Flu-like symptoms and muscle aches in the legs and buttocks.  These prodomal symptoms can tip-off sufferers of an impending eruption.

Compare and contrast herpes encephalitis and herpes meningitis.

 Encephalitis                                                        Meningitis

From recurrent HSV 1 infection in adults     10% of primary HSV 2 infections adl

    From primary HSV 2 infection in neonates

    Headache, irritability, seizures                      Neck stiffness, headache, vomiting

Severe, must be treated promptly with ACV      Usually resolves in one week

Summarize major features of neonatal herpes infections.  What are the risks of infection to the neonate if the mother has a primary infection? A recurrent infection?

      Symptoms usually develop within the first two weeks post-partum as 95% of infections occur during delivery.  Infection may be limited to the skin, eyes, or mouth, and these patients typically exhibit a “zoster-form” rash.  More severe infections are disseminated and can involve multiple organs and the CNS (encephalitis).  25% of severely infected neonates will die even with treatment, and another 50% will suffer significant sequelae.  The risk to a newborn if the mother is undergoing a primary infection is nearly 30%, but because maternal antibodies are protective, the risk drops to 2-3% if the mother is undergoing a recurrent infection.  Pregnant women with a history of genital recurrences should be given acyclovir in the final trimester of pregnancy to suppress reactivation.

What is the incidence of HSV 1 and 2 infections in the US population?

HSV 1: up to 60% of US adults

      HSV 2: 10% in young adults and adult males; 20% in adult females

How is genital herpes diagnosed? Herpes encephalitis?

Genital herpes diagnosis: Viral culture from swabs of vesicles and/or fluorescent antibody tests; in the absence of lesions, serology can determine past infections

      Herpes encephalitis diagnosis: PCR amplification of HSV genome fragments from CSF

             Acyclovir is more active in infected cells because it is dependent on the herpes thymidine kinase to obtain the first of three phosphate groups necessary to become an active nucleoside analog.  Once acyclovir achieves its triphosphate form, it becomes a preferred substrate of the herpes DNA polymerase and is incorporated into replicating virus genome DNA strands where it prevents further elongation.

How is acyclovir (or famciclovir) used to control genital infections? To treat encephalitis?

Treatment of genital infections: lose dose oral ACV or single high dose of famciclovir; daily low dose suppressive therapy for those with frequent recurrences

      Treatment of encephalitis: IV, high dose ACV for up to 3 weeks at first suspicion of illness

 How does foscarnet (phosphonoformate) differ from acyclovir?

.      Foscarnet is a pyrophosphate analog, not a nucleoside analog.  It jams the pyrophosphate exchange site in the viral DNA polymerase, blocking DNA replication.  Significantly, it does not need phosphorylation to become active, and therefore is effective against viruses that have mutations in the thymidine kinase gene that renders them resistant to acyclovir.

 How does Abreva (docosanol) work?