Virology

VARICELLA-ZOSTER VIRUS( VZV )(HHV3)

Introduction
Varicella - zoster virus (VZV) Virus causes two different diseases. I. Varicella (chickenpox) Acute mild highly contagious disease, occur in epidemics, among children result from primary infection with the virus. Latent in sensory ganglion. II. Zoster (shingles) Response of the partially immune host to reactivation of varicella virus present in a latent form in sensory ganglion result in zoster. Sporadic disease of adults. Characterized by vesicular cutaneous rash limited in distribution to part of the skin innervated by a single sensory ganglion.

Properties of the virus

Pathogenensis
Source Patients with varicella or zoster lesion (highly contagious). Mode of transmission Respiratory droplet. Route of entry Mucosa of upper respiratory tract or conjunctiva. Replication Replicate first locally in the regional lymph node. Spread After local replication in the regional LN, the virus circulate in the blood ( primary viremia), to invade & replicate in the liver & spleen, followed by secondary viremia. Lastly the virus localize in the skin & mucous membrane (target organ). Lesion in target organ Skin & mucosal lesions initiated by viral infection of capillary endothelial cells & epidermal cells, result in swelling of epithelial cells, ballooning with accumulation of tissue fluid (vesicle formation). Sever form varicella Occur in neonates, or IC. patients, here varicella lesion can occur in other organ like liver & lungs with multinucleated giant cells. Viral shedding Respiratory secretions. Latent The virus become latent in the dorsal root sensory ganglion.

Reactivation Trigger factor is weaning of immunity, allows viral replication to occur in dorsal root ganglion result in 1- Acute inflammation in the sensory nerves & ganglia. 2- Distribution of lesion in the skin (vesicles) corresponds closely to the area of innervation from an individual dorsal ganglion ( dermatomal distribution).

Clinical findings

I. Varicella ( chickenpox) Mild disease among children. Sever disease occur in immunocompromised patients & neonates. Incubation period is 10-21 days. Subclinical varicella is unusual. Characterized by fever, generalized vesicular eruption of skin & mucous membranes. Characteristics of rash: vesicular, generalized on the trunk, then on the face, limbs, buccal & pharyngeal mucosa in the mouth. All stage of skin lesion macules, papules, vesicles & crusts seen at one time. The rash lasts about 5 days, & most children develop several 100s skin lesions. Complications Rare in normal children, include 1- Encephalitis especially in neonatel varicella (limited immunity), mortality 30%. 2- Varicella pneumonia most common complication in neonates, adults, immunocompromised patients, with primary infection.

II. Zoster (shingles) Usually occur in immunocompromised, but occasionally develops in healthy adults. Starts with sever pain in the area of skin or mucosa supplied by one or more groups of sensory nerves & ganglia (often only a single ganglion involved). The trunk, head,& neck are most commonly affected, with ophthalmic branch of trigeminal nerve involved in 10-15% of cases. Within a few days a crop of vesicles appears over the skin supplied by the affected nerves (dermatomal distribution). Complications 1- Post-herpetic neuralgia Common, protracted pain that may continue for months. 2- VZV pneumonia Responsible for deaths that occur in immunosuppressed patients with zoster (< 1% of patients).

Chicken pox

Shingles

Laboratory Diagnosis

Diagnosis is clear from clinical presentations. Samples: vesicle fluid & scrapings form the base of skin lesions. I. Direct Methods 1. Cytology Smears from base of the lesions will reveal characteristic multinucleate giant cells ( Tzanck smear). 2. Electron microscopy Herpesviruses can be recognized by the morphologic appearance of the viral particles in vesicular fluid examined by EM. 3. Immunofluorescence cytology Smears from the lesions can be examined by immunofluorescence to detect VZV antigen. 4. PCR PCR assays for VZV available & have been used in the diagnosis of VZV encephalitis from CSF specimens.

II. Virus isolation Type of cell culture is human fibroblasts, for the identification of VZV replication: 1- CPE produced by VZV is produced within 3-7 days, consists of a typical ovoid focal pattern. 2- Immunofluorescence of the cell sheet by monoclonal antibodies is the method of choice for identification.

III. Serology 1- Specific VZV IgG in paired acute & convalescent sera. 2- Specific VZV- IgM. Techniques Complement fixation test (CFT) most frequently used test. It is perfectly adequate for the diagnostic purposes . Immunofluorescence test (IF) more sensitive than CFT.  Enzyme immnuoassay (EIA) These are the most sensitive methods available & therefore are the preferred method.

Epidemiology

Varicella is highly communicable, common epidemic disease of childhood (mostly in children < 10 years), adult cases do occur. More common in winter & spring. Spread airborne droplets. Patients are infectious shortly before the appearance of vesicles to about 5 days later. Zoster occur sporadically, chiefly in adults without seasonal prevalence. Prevention 1- A live attenuated varicella vaccine was approved in 1995 for general use. The vaccine is highly effective in children (80-85%), but less in adults( 70%). Isolation of patient with varicella from exposed to IC patients & neonates, because of serious squeals of pneumonia, encephalitis & death. IC. patients exposed to varicella, VZV.Ig & acyclovir administration used to modify the disease (prophylactic use).


Treatment: 1- Varicella in normal children is mild disease & require no treatment. 2- Sever varicella ( neonates, immunocompromised patients ) is potentially fatal that require treatment 3- Varicella zoster immune globulin used to prevent the development of the illness, & has no therapeutic value once the varicella has started. 4- Infants born to women with peripartum chickenpox (from 5 days before delivery), VZIG to infant reduces severity & mortality of neonatal infection. 5- Antiviral compound shown to be effective including: Acyclovir ( treatment of choice). Famciclovir. Valacyclovir. Foscarnet.

Epstein-Barr virus

(EBV) (HHV4)

Introduction

Epstein Barr virus (herpes virus) is a causative agent: 1- Primary infection: result in a disease called Acute infectious mononucleosis (heterophile positive disease). mononucleosis mean polyclonal stimulation of B lymphocytes. 2- Latent: B Lymphocytes. 3- Re-activation: EBV is a factor in development of Naso-pharyngeal Carcinoma. Burkitts Lymphoma.


Properties of the virus
1- EBV have the general properties of herpesviruses (spherical, large size, enveloped, with icosahedral nucleocapsid, DNA virus). 2- Genome is DNA, contain 172 kbp . 3- Viral antigens: Latent antigen: (produced by latently infected B cells, include EBNAs, and LMPs, there presence indicate that EBV genome is present). Early antigen: (non structural protein, it indicate the onset of productive viral replication). Late antigen: (Structural protein and viral capsid antigen and viral envelope glycoproteins). 4- Target cell for EBV is the B lymphocyte.

Pathogenesis of Infectious Mononucleosis

Source: Human with viral excretion in saliva, patients shed low levels of virus for weeks to months after infection. Mode of transmission: EB virus is transmitted by infected saliva & respiratory droplets. (kissing disease). Initial viral replication occur in epithelial cells of the oropharynx or surface B lymphocytes, & salivary glands. Interaction of EBV virus with cellular receptor: The EBV initiate infection by binding to viral receptors on B cells (CR2 or CD21). Spread: virus is spread by blood stream & disseminated through the body to liver , spleen, & lymph nodes. The proliferation & expansion of EBV infected cells with activated T-cells result in enlargement of these organ. Most people shed the virus in saliva for weeks to months after infection.

Latent phase: Latent site lymphoid cells, small number of infected lymphocytes persist for lifetime of the host (one in 106 of B cells ). Reactivation of EBV: evident by Increased of virus in saliva & usually is clinically silent. Reactivate infection in immunocompromised (immunosuppressive drugs, or diseases) carries serious consequences. The EBV infected B lymphocytes express differentiated functions : 1- Secretion of immunoglobulin: IgA & IgG are common, whereas IgM synthesis is rare. Auto antibodies are typical of the disease, like rheumatoid factor, antinuclear antibodies. Heterophil antibodies that react with antigen on sheep erythrocytes. 2- Expressed B-cell activation products e.g CD23. 3- Expression of viral gene in the infected cells including six different EBV antigens ( EBNA1-6) and two latent membrane proteins (LMP1, LMP2). 4- Few infected B-cells release virus particles (<10%).

Clinical findings and diseases

Primary infection in children are asymptomatic, in adolescent & young adults the classic disease with primary infection is infectious mononucleosis. Infectious mononucleosis (IM): Incubation period is 30-50 days. Symptoms: headache, fever, malaise, fatigue & sore throat. Enlarged lymph node & splenomegaly are characteristic. Maculopapular rash especially after ampicillin therapy. Some patients develop sign of hepatitis. The illness last for 2-4 weeks During the illness there is increase lymphocytes in the blood, with large atypical lymphocytes. Complication is rare in normal hosts, like hepatitis.

Laboratory diagnosis

1- Nucleic acid hybridization: the most sensitive means of detecting EBV in patients respiratory secretion, blood. 2- Serology: 2 types of antibodies are developed in the serum:

I. Specific EBV antibodies (EBV specific IgG, or EBV specific IgM), detected by serological procedures like ELISA test, indirect immunofluorescent test using EBV-positive lymphoid cells. The presence of EBV IgM type is indicative of current infection. Antibody of IgG type is a marker of past infection & indicate immunity. II- Less specific EBV antibodies is called heterophil antibodies that agglutinate sheep cells by a test called heterophil agglutination test (Paul-Bunnell test). Most patients develop these antibodies in the serum during active infection. 3- Isolation EBV: Saliva, Blood inoculated into cell culture containing normal human lymphocytes obtained from umbilical cord blood. Virus need 6-8 weeks to grow.

Prevention and Treatment There is no EBV vaccine available. Acyclovir reduces EBV shedding from oropharynx during period of drug administration but it does not affect the symptoms of IM.

( Cytomegalovirus )(CMV)(HHV5)

Introduction

CMV are common cause of human disease. The name for the virus is derived from massive enlargement of CMV infected cells. Properities of the virus: 1- General properties of herpes viruses. 2- Genome : DNA, size 240 kpb, largest genome among human herpesviruses. 3- Infected cell surface glycoprotein act as receptor for FC portion of Ig, so Ig bind non specifically to the cells surface and provide protective coat to help the virus to evade immune elimination.

Pathogenesis:

Human is the source for infection. Transmission need close person to person contact, by close contact with virus bearing materials (saliva, blood, genital secretion). The virus cause systemic infection involve monocytes, T & B cells, liver, lung, and kidneys. Result in infectious mononucleosis like syndrome in normal hosts, in IC. patients result in sever disease, disseminated infection with pneumonia. Virus shedding from pharynx, urine for months or years after primary infection. Lifelong latency in lymphocytes. Reactivation: asymptomatic viral shedding in normal host, or serious involvement of lungs (CMV pneumonia) in IC hosts.

(Diseases caused by the CMV)Depend on the age and immune status of the patient

1- Fetal infection: Death, or born with congenital anomalies (high frequency 10% caused by CMV), result from intrauterine infection from mother mostly with primary infection. 2- Newborn baby: disease called generalized cytomegalic inclusion disease (CNS & reticuloendothelial involvement) result in mental retardation & hepatosplenomegaly). Caused by intrauterine (mostly from primary maternal infection) or early postnatal infection with the CMV (genital tract of mother or breast milk). 3- Adolescence: Inapparent infection, when symptomatic it result in infectious mononucleosis (heterophil - negative), clinically characterized by fever, malaise, lymph node enlargement, liver function abnormalities & lymphocytosis with atypical mononuclear cells, Paul-Bunnel test negative. 4- Adults: sever CMV pneumonia are frequently found in IC adults, usually fatal.

Laboratory diagnosis

1- Detection of CMV nucleic acid using Polymerase Chain Reaction (PCR) : Commonly used for routine diagnosis of CMV diseases. PCR designed to detect replicating virus not latent virus genome.2- Detection of CMV antigen using immunoflourescent test (IF) : Viral antigen can be detected in peripheral leukocytes using monoclonal antibodies against viral antigens using IF testy. 3- Detection of specific CMV antibodies using Enzyme Linked Immunosorbent assay (ELISA) : I. Specific CMV-IgG which indicate past infection, rising titer indicate recent infection. II. Detection of viral IgM antibodies suggest current infection.4- Isolation of the CMV: Human fibroblast, throat wash & urine used, 2 – 3 weeks needed for CPE to appear as multinucleated giant cells, or immunoflouresent test to detect viral antigen in cell culture.

Treatment: Gancyclovir is used to treat life threatening CMV infection in IC. Patients, CMV pneumonia .