B52 Cytomegalovirus , Mumps.pdf

IA Session: Cytomegalovirus & Mumps Infection Dr. Anita Devi Krishnan Thantry Department of Microbiology Faculty of Medicine [email protected] 04th July 2024 10.30 am – 12.30 pm Learning Outcomes: At the end of the session, the student should be able...

IA Session: Cytomegalovirus & Mumps Infection Dr. Anita Devi Krishnan Thantry Department of Microbiology Faculty of Medicine [email protected] 04th July 2024 10.30 am – 12.30 pm Learning Outcomes: At the end of the session, the student should be able to: Cytomegalovirus: Describe the general properties of the causative agent C2 Describe the transmission, pathogenesis, clinical features and complications C2 Describe the laboratory diagnosis C2 Mumps virus : Describe the general properties of the causative agent C2 Describe the transmission, pathogenesis, clinical features, and complications of the causative agent C2 Describe the laboratory diagnosis C2 State the preventive measures C1 2 Cytomegalovirus Herpes virus family → beta subfamily. Worldwide occurrence. LATENCY is a hall mark Structure: Spherical with icosahedral symmetry of capsid. Capsid : 162 hollow hexagonal and pentagonal capsomeres Nucleocapsid: Electron-dense core Double-stranded DNA genome 3 Cytomegalovirus ( Structure Contd.) Envelope: Lipoprotein, trilaminar Lipid part derived from the nuclear membrane of the infected host cell. Has viral glycoproteins spikes which bind to specific host receptor and mediate virus entry. gB : major surface glycoprotein →Binding and entry into host cell and cell to cell transmission. gH : Target for complement independent neutralizing antibodies. Tegument : Outer to capsid, amorphous proteinaceous layer VP16 enzyme : Redirect host cellular proteins and enzymes to involve in viral nucleic acid replication VHS (Virion Host Shutoff) protein: Shuts off host cell protein synthesis. 4 Transmission of CMV Commonest: Person to person via close contact with an individual who is excreting the virus ( saliva, urine , other body fluids) esp., young children. Other modes of transmission: Placenta → congenital CMV Blood transfusions Organ transplantation Breast milk -rare Sexual transmission Individuals at an increased risk for CMV infection: Attend or work at daycare centers Blood transfusions Multiple sex partners Recipients of CMV mismatched organ or bone marrow transplants. Manipal Patients deficient in cell-mediated immunity are at greatest risk for CMV disease. University College Malaysia 5 Replication of Human Cytomegalovirus Attachment to host cells : Fusion of the viral glycoproteins to host receptors Entry : Fusion mediates endocytosis of the virus into the host cell. Uncoating Nuclear translocation: Capsid is transported to the nuclear pore Viral DNA is released into the nucleus. Replication in nucleus: Regulated by tegument proteins Initiate expression of replication genes: Viral immediate-early (IE) gene → Delayed early (DE) genes → initiate viral genome replication → Late (L) gene → initiates capsid assembly in the nucleus 6 Replication of Human Cytomegalovirus Egress from nucleus to cytosol. Trafficked to the viral assembly complex (AC) = endoplasmic reticulum (ER), Golgi apparatus + endosomal machinery Assembly : Viral DNA & proteins assemble in nuclear viral factories and bud through inner lamella of the Capsids associate with tegument proteins in the cytosol nuclear membrane Budding: At assembly complex, capsids acquire tegument and viral envelope with insertion of glycoproteins by budding into intracellular vesicles Release : Exocytosis at the plasma membrane 7 Life cycle of Cytomegalovirus 8 Pathogenesis of Human Cytomegalovirus Site of multiplication: Epithelial cells of gland & mucosal tissue, smooth muscle cells, fibroblasts, macrophages, dendritic cells, hepatocytes, and vascular endothelial cells. Broad tropism for cells → facilitates spread systemically and inter-host spread Primary replication : Mucosal epithelium (direct contact with infectious secretions from an infected individual) Prolonged secretion of virus in saliva for months Dissemination via leukocyte → viremia → systemic spread (last for months) Latency : Myeloid cells of the bone marrow → life-long infection with sporadic reactivation. Salivary glands Reactivation: Excretion in saliva, semen, cervical and vaginal secretions, and breast milk 9 Pathophysiology of Latency Host immune response following primary human cytomegalovirus (HCMV) infection is effective at stopping virus replication and dissemination. But virus is never cleared by the host. Persistence due to ability of HCMV to establish latency in myeloid cells and circulating monocytes Primary infection& resolution: HCMV replicates and disseminates during which time the host generates an Latency & reactivation: HCMV replicates and disseminates leading to effective immune response which includes natural killer infection of myeloid progenitors and the establishment of latent cells, neutralizing antibodies and a high frequency of CD4+ infection in e.g., CD34+ bone marrow progenitor cells. Reactivation of and CD8+ T cells. This controls viral replication and resolves virus from these sites followed by new virus replication and the primary Manipal University College Malaysia infection. productive replication induces secondary immune responses..10 Pathophysiology CMV is a lytic virus that causes a cytopathic effect Enlarged cell with viral intranuclear basophilic inclusion bodies with an "owl's eye“ appearance →pathologic hallmark of CMV infection CMV is an immunomodulatory virus may aggravate underlying immune disorders. CMV harbors the largest number of genes dedicated to evading innate and adaptive immunity in the host. CMV-specific CD4+ and CD8+ lymphocytes play an important role in immune protection after primary infection or reactivation of latent disease. Manipal University College Malaysia 11  Pathogenesis of Human Cytomegalovirus Depends on immune status in adults: Immunocompetent : Asymptomatic/ Mild &self limiting ( progress to latency is many) Immunocompromised: Life-threatening risk in (organ transplant recipients ,HIV ) In pregnant women: Leading infectious cause of congenital neurological disease due to transplacental transmission. Importance of CMV antibodies: CMV specific IgM antibodies: Seen in primary infection and persist for 3 or 4 months Initially low avidity IgM are produced, later convert to high avidity IgM NOT PRODUCED IN RECURRENT INFECTIONS IN IMMUNOCOMPETENT INDIVIDUALS. CMV IgG antibodies : Produced at time of primary infection and PERSIST LIFELONG. In intrauterine infections: both IgM and IgG are produced by the fetus Cell-mediated immunity (CMI) : Key role in the suppression of CMV infection. 12 Pathogenesis of CMV infection 13 Clinical manifestations Immunocompetent: Asymptomatic in many Acute CMV Infection/ “CMV mononucleosis”: Fever, malaise, enlarged lymph nodes, sore throat, muscle aches, loss of appetite, enlarged liver or spleen, and fatigue. Hepatitis-like symptoms and signs may include appetite loss, yellow eyes, nausea, and diarrhea Mimic infectious mononucleosis caused by Epstein-Barr virus or liver infection by hepatitis A, B, or C. Immunosuppressed: CMV retinitis, pneumonia, esophagitis, colitis, hepatitis, encephalitis (behavioral changes, seizures, or coma). 14 Clinical manifestations Congenital : No symptoms at birth in 90% Symptomatic in 10% Features of congenital CMV infection: CNS abnormalities – microcephaly, mental retardation, spasticity, epilepsy, periventricular calcification Choroidoretinitis and optic atrophy Sensorineural deafness ( Major finding – 20% cases) Hepatosplenomegaly and jaundice which is due to hepatitis Pneumonitis Myocarditis Thrombocytopenic purpura Hemolytic anemia Late sequelae – damage to the enamel forming organ of the teeth resulting in yellow discoloration of the teeth and brittleness. 15 Clinical Manifestations Rare Manifestations: Causes mucoepidermoid carcinoma ? Responsible for prostate cancer Associated with acute graft versus host disease (GVHD) in bone marrow transplant recipients 16 Laboratory diagnosis Specimens: Urine, saline, tissue biopsy, other body fluids Cytopathology- Compound Microscopy : Biopsy material Hematoxylin-eosin–stained lung section showing typical owl-eye inclusions (480X). Histopathological diagnosis is the “gold standard” Warde Medical Laboratory, Ann Arbor, Michigan. Stains: Giemsa, Wright, hematoxylin-eosin, Papanicolaou “Owl’s eyes appearance”: Intracellular cytomegalic inclusions surrounded by clear halo Electron Microscopy Virions in the urine of congenitally infected infants Virus isolation Human embryo lung fibroblasts / HELA cell lines Produces a typical focal cytopathic effect 17 Laboratory diagnosis Immunohistochemistry/Tissue immunofluorescence : Samples: Tissue(liver, lung) or body fluid, frozen sections of biopsy tissue Infected lung and liver cells stained by specific anti- CMV antibodies to locate CMV antigens Visualized by fluorescently labeled antibodies or enzyme-labeled secondary antibodies Stained slides are examined by fluorescent or light microscopy. 18 Laboratory Diagnosis CMV pp65-antigen-positive granulocyte Antigen detection CMV pp65 antigen assay in leukocytes Detects messenger matrix proteins of CMV Serology Tests: Complement fixation, Enzyme-linked immunosorbent assay (ELISA), Anti- complement immunofluorescence, Radioimmunoassay, and Indirect hemagglutination. Acute or recent infection: IgM antibodies (IgM capture assays ) Past infection: CMV IgG. 19 Laboratory Methods Molecular methods Polymerase chain reaction (PCR) : Rapid and sensitive method → amplification of nucleic acids. Qualitative or quantitative Quantitative Real-Time PCR: (amount of viral DNA is measured). Used for continuous monitoring of immunocompromised individuals To identify patients at risk for CMV disease for preemptive therapy To determine response to treatment. Nucleic acid sequence-based amplification (NASBA) : Sequence-based amplification of pp67 viral mRNA in a background of DNA using specific isothermal technique of amplification. 20 Laboratory Diagnosis Typical owl-eye inclusion bodies of Cytomegalovirus with enlarged cells (“cyto-megalo”) Cowdry Type B basophilic intranuclear inclusion bodies 21  Laboratory Diagnosis IgG avidity assays: Helps distinguish primary CMV infection from a past infection. Measures binding strength between IgG antibodies and virus Principle: In primary CMV infection, IgG antibodies initially have low binding strength (low avidity) Over 2 to 4 months they mature to high binding strength (high avidity). Congenital CMV : Low avidity indicates an increased risk whereas high avidity indicates a low risk of congenital infection, 22 Treatment Ganciclovir and foscarnet → life-threatening CMV infection in immunocompromised patients. Acyclovir and valacyclovir → bone marrow and renal transplant patients. Foscarnet and cidofovir are used as second-line drugs. Prevention and control Isolation of newborns with generalized cytomegalic inclusion disease. Screening of transplant donors and recipients for CMV antibody. Human IgG prophylaxis in transplant recipients. Live and recombinant vaccines are under development. 23  Mumps Pleomorphic (100 to 600 nm) Spherical Nucleocapsid : Ribonucleoprotein complex → hollow tube with a central core RNA genome : negative-sense, unsegmented ,single-stranded Associated proteins: Large (L) protein :RNA polymerase Polymerase phosphoprotein (P) protein: Facilitates RNA synthesis Nucleoprotein (NP) protein: Helps maintain genomic structure. Matrix (M) protein: Lines the inside of the virion envelope Envelope: Contains two glycoproteins: Hemagglutinin-neuraminidase [HN] / hemagglutinin [H]/ glycoprotein [G] protein: Attachment protein Fusion (F) protein: Promotes fusion of the viral and host cell membranes 24 Epidemiology of Mumps Virus Endemic worldwide. Vaccine preventable disease : Live virus vaccine Highest incidence : children aged 5–9 years. Outbreaks still being reported across the world ( U.S; UK). Crowding favors dissemination of the virus. Commonest presentation: PAROTITIS