RNA Viruses PDF

Untung Yuli Prastiawan (aka Wawan Geni) (1982–). The Little Time, 2017 (detail). Burning techniques applied to canvas, 35.4 in × 27.6 in/90 cm × 70 cm. Wawan Geni Collection. Digital image courtesy of Wawan Geni. RNA VIRUSES, PRIONS TOPIC VIROLOGY SUB...

Untung Yuli Prastiawan (aka Wawan Geni) (1982–). The Little Time, 2017 (detail). Burning techniques applied to canvas, 35.4 in × 27.6 in/90 cm × 70 cm. Wawan Geni Collection. Digital image courtesy of Wawan Geni. RNA VIRUSES, PRIONS TOPIC VIROLOGY SUBJECT MICROBIOLOGY 2 INST. RNA Viruses and Prions Topic Outline Orthomyxoviruses Bunyaviruses Arenaviruses Paramyxoviruses Rhabdoviruses Flaviviruses Coronaviruses Retroviruses Picornaviruses Reoviruses Prions and Spongiform Encephalopathies 2 RNA Viruses Overview ©The McGraw-Hill Companies, Inc. 3 FAMILY ORTHOMYXOVIRIDAE Influenza viruses Properties ๏ Virion: spherical pleomorphic (helical nucleocapsid) ๏ Genome: ssRNA segmented, (-)sense ๏ Proteins: 9 structural proteins, 1 nonstructural proteins ๏ Envelope: HA (hemagglutinin), NA (neuraminidase) ๏ Replication: Nuclear transcription Notable characteristics ๏ Genetic reassortment common among members of the same genus. ๏ Influenza viruses cause worldwide epidemics FAMILY ORTHOMYXOVIRIDAE Influenza viruses Electron micrograph if influenza virus A/Hong Kong/1/68/(H3N2) ๏ Note pleomorphic shapes and glycoprotein projections covering particle surfaces (315,000x) (Courtesy of FA Murphy and EL Palmer) glycoprotein envelope FAMILY ORTHOMYXOVIRIDAE Influenza viruses Three distinct influenza virus types: Amost common, B, C Viral infection ๏ Virus attaches to, and multiplies in, the cells of the respiratory tract ๏ Segments of RNA genome enter the nucleus (transcribed then translated) ๏ Synthesized viruses assembled and release by budding occurs at the plasma membrane FAMILY ORTHOMYXOVIRIDAE Influenza viruses: Viral cycle Receptor mediated endocytosis Replication Transcription Translation (Budding) glycoprotein spikes synthesized on the host membrane ©The McGraw-Hill Companies, Inc. FAMILY ORTHOMYXOVIRIDAE Influenza viruses: The glycoprotein spikes Hemagglutinin (H)/HA ๏ Major Ag that neutralises host’s Ab— used to anchor virus to host cell receptors ๏ Agglutinates erythrocytes under certain conditions ๏ Variability is responsible for continual evolution of new strains and subsequent influenza epidemics ๏ 15 subtypes Neuraminidase (N)/NA ๏ Tetramer that hydrolyses mucus and assists in viral budding and release ๏ Helps prevent self-aggregation of virions by removing sialic acid (sialidase enzyme) residues from viral glycoproteins FAMILY ORTHOMYXOVIRIDAE Influenza viruses Type H/H Subtype Strain/History A H1N1 Spanish flu pandemic of 1918 A/New Jersey/76 (swine flu outbreak) A/USSR/90/77 A/Texas/36/91 H2N2 A/Singapore/57 (Asian flu pandemic) A/Japan/62 A/Taiwan/64 H3N2 A/Hongkong/68 (Hong Kong flu pandemic) A/Johannesburg/33/94 B None B/Harbin/07/94 C None C/JHB/2/66 + The last number is the year the virus appeared FAMILY ORTHOMYXOVIRIDAE Influenza viruses: The Genetic reassortment Influenza viruses undergo frequent antigenic changes in HA and NA, a selective advantage over the parental virus in the presence of Ab detected against the original strain. Antigenic Drift ๏ Due to accumulation of point mutations in the gene resulting to AA changes in the protein— alters the antigenic sites. ๏ Influenza B— only undergo Ag drift Antigenic Shift ๏ Reflects changes in the sequence of a viral surface protein— too extreme to be explained by mutation. ๏ One of the genes or RNA strands is substituted with a gene strand from other influenza virus from a different animal host—genetic reassortment between human and avian viruses FAMILY ORTHOMYXOVIRIDAE Influenza viruses: Antigenic Shift FAMILY ORTHOMYXOVIRIDAE Influenza viruses Influenza A ๏ Acute highly contagious respiratory illness ๏ Seasonal, pandemics; among top 10 causes of death in U.S.— most commonly among elderly and small children ๏ Binds ciliated cells of respiratory mucosa ๏ Causes rapid shedding of cells, stripping the respiratory epithelium; severe inflammation ๏ S|S: fever, headache, mylagia, pharyngeal pain, shortness of breath, coughing ๏ Weakened host defense predispose patients to secondary bacterial infections, especially pneumonia. FAMILY ORTHOMYXOVIRIDAE Influenza viruses: Diagnosis, Treatment, & Prevention Rapid immunofluorescence ๏ Detects Ag in pharyngeal specimen; serological testing to screen for Ab titer Treatment ๏ Control symptoms; Oseltamivir (Tamiflu), Zanamivir (Relenza), rimantadine, and amantadine; Adamantium & Vibranium ๏ High rate resistance: Amantadine & Rimantadine Annual trivalent (quadrivalent ⇥ Pentavalent) vaccine recommended This Slide is Intentionally Left Blank ! BUNYAVIRUSES Introduction Properties ๏ Virion: spherical ๏ Genome: triple-segmented, (-) sense or ambisense (+/-) ssRNA ๏ Proteins: 4 major polypeptides ๏ Envelope: yes ๏ Replication: cytoplasm ๏ Assembly: budding into the Golgi Notable characteristics ๏ The virion contains a transcriptase BUNYAVIRUSES Overview: Hanta virus Transmitted zoonotically and causes periodic epidemics that are extremely dangerous— BSL 4 classified Transmitted by insects and ticks ๏ California encephalitis, Rift Valley fever, Korean hemorrhagic fever ๏ American bunyavirus is a hanta virus, Sin Nombre— emerging disease Carried by deer and harvest mice Transmitted via airborne dried animal waste This Slide is Intentionally Left Blank ! ARENAVIRUSES Introduction Properties ๏ Virion: spherical ๏ Genome: ssRNA, double segmented, (-)sense and ambisense (+/-) ๏ Proteins: 4 major polypeptides ๏ Envelope: yes ๏ Replication: cytoplasm ๏ Assembly: incorporate ribosomes and bud from plasma membrane Notable characteristics ๏ The virion contains a transcriptase ARENAVIRUSES Overview Lassa fever (Lassa virus); Argentine hemorrhagic fever (formerly Junin virus); Bolivian hemorrhagic fever (Machupo virus | aka black typhus), and; Lymphocytic choriomeningitis Closely associated with rodent host Transmission through aerosols and contact Divided into two groups: Old world New world This Slide is Intentionally Left Blank ! PARAMYXOVIRUSES Introduction Properties ๏ Virion: spherical, pleomorphic (helical nucleocapsid) ๏ Genome: ssRNA linear, nonsegmented, (-)sense ๏ Proteins: 6-8 nonstructural proteins ๏ Envelope: viral glycoprotein (G, H, of HN)— sometimes carries HA or NA activity; fusion (F) glycoprotein ๏ Replication: cytoplasm, particles bud from plasma membrane Notable characteristics ๏ Antigenically stable ๏ Particles are labile yet highly infectious! PARAMYXOVIRUSES Introduction Parainfluenza, mumps virus Morbilivirus, measles virus Pneumovirus, respiratory syncytial virus ๏ Respiratory transmission ๏ Envelope has glycoprotein and F-spikes that initiate cell-to-cell fusion ๏ Fusion with neighbouring cells— syncytium or multinucleate giant cells form. ๏ Prevention: PARAMYXOVIRUSES Subfamilies and their notable characteristics Paramyxovirinae Pneumovirinae Metapneumo- Respirovirus Rubulavirus Morbillivirus Henivirus Pneumovirus virus Mumps, Hendra, Nipah Respiratory Heman meta- Human viruses Parainfluenza Measles Parainfluenza (zoonotic) syncytial virus pneumovirus F glycoprotein + + + + + + Hemolysin + + + ~ 0 0 Hemagglutinin + + + 0 0 0 Headsorption + + + 0 0 0 Neuraminidase + + 0 0 0 0 PARAMYXOVIRUSES Viral life cycle Attachment [HN, H, or G] [CD46 o rCD150] Replication Cytoplasm Transcription Translation Budding PARAMYXOVIRUSES Parainfluenza HPIV seasons PARAMYXOVIRUSES Mumps Epidemic parotitis, self-limited, associated with painful swelling of parotid salivary glands Human are the only reservoir 40% of infections are subclinical, long-term immunity Incubation 2-3 weeks fever, muscle pain and malaise, classic swelling of one or both cheeks Usu. uncomplicated invasion of other organs; in 20-30% of infected adult males, epididymis and testes become infected— sterility is rare Live attenuated vaccine MMR PARAMYXOVIRUSES Mumps Coughing, sneezing, or taking Sharing items that may have saliva on them, such as water bottles or cups Participating in close-contact activities with others, such as playing sports, dancing, or kissing Touching objects or surfaces with unwashed hands that are then touched by others. PARAMYXOVIRUSES Measles (Rubeola) Acute, highly infectious characterized by fever, respiratory symptoms, and maculopapular rash Introduction of live-virus vaccine reduced prevalence Humans— the only natural host Virus enters via the URT and multiplies locally → regional lymphoid tissue where further multiplication occurs Stages of infection: Viral invasion Prodromal phase Rash— interaction of T cells with virus-infected cells in the small blood vessels. For defective cell-mediated immunity— no rash develops Viral clearance— coincident with fading of rash PARAMYXOVIRUSES Measles (Rubeola) Replication begins in the RT epithelium ⇥ monocyte-macrophages endothelial cells, ⇥ and epithelial cells in ๏ blood, spleen, lymph nodes, lung, thymus, liver, and skin, and to the mucosal surface of GIT, RT, GUT. ๏ SSPE: Subacute sclerosing panencephalitis PARAMYXOVIRUSES Measles: SSPE Subacute sclerosing panecephalitis— most serious complication Progressive neurological degeneration of the cerebral cortex, white matter, and brain stem. ๏ 1 case in a million infections ๏ Involves a defective virus spreading through the brain by cell fusion and destroys cells ๏ Leads to coma and death in months or years Prevention ๏ Attenuated viral vaccine— MMR This Slide is Intentionally Left Blank ! RHABDOVIRUSES Introduction Properties ๏ Virion: bullet-shaped ๏ Genome: ssRNA linear, nonsegmented, (-)sense ๏ Proteins: 5 major proteins; 1 is enveloped glycoprotein ๏ Envelope: present ๏ Replication: cytoplasm, particles bud from plasma membrane Notable characteristics ๏ Wide array of viruses with broad host range ๏ Group includes the deadly rabies virus (Genus Lyssavirus) ๏ Primary reservoir are wild mammals RHABDOVIRUSES Illustrations Glycoprotein spikes Matrix protein Nucleocapsid ๏ Electron micrograph (100,000x) ๏ Schematic model ๏ Shown a vesicular stomatitis virus negatively ๏ Virus showing glycoprotein spikes extending from stained with potassium phosphotungstate. ℹRM lipid envelope that surrounds internal nucleocapsid McCombs, M Benyesh-Melnick, JP Brunschwig & the matrix protein lining the envelope RHABDOVIRUSES Dissemination to extra CNS sites eg., salivary glands via cranial nerves Pathogenesis: Rabies Virus -laden saliva contaminates tissue Infection of neurons Infection of spinal neurons & Axonal transport to brain Replication within ventral horn Rabies virus neurons and dorsal root ganglia Skeletal Muscle cell Viral entry into skeletal muscle cells and replication Retrograde axonal transport to spinal cord Axonal terminal Budding into neuromuscular junction and uptake into peripheral nerve endings. Virus can also enter peripheral nerves without muscle involvement RHABDOVIRUSES Pathogenesis: Rabies Clinical phases: ๏ Prodromal phase Fever, nausea, vomiting, headache, fatigue Some experience pain, burning, tingling sensations at site of wound. ๏ Furious phase Agitation, disorientation, seizure, twitching, hydrophobia ๏ Dumb phase Paralysed, disoriented, stuporous ๏ Progression to coma → death RHABDOVIRUSES Diagnosis Immunofluorescence or immunoperoxidase staining of tissue biopsy ๏ Use of antirabies monoclonal Ab— rapid! Presence of Negri bodies in the brain or spinal cord ๏ Definitive pathologic diagnosis ๏ Sharply demarcated, ~spherical (2-10um) diameter, with distinct internal structure with basophilic granules in an eosinophilic matrix. RT-PCR Viral isolation Serology (Nt test) Animal observation This Slide is Intentionally Left Blank ! FLAVIVIRUSES Introduction Properties ๏ Virion: spherical ๏ Genome: ssRNA linear, (+)sense ๏ Envelope: 3 structural polypeptides, 2 glycosylated ๏ Replication: cytoplasm ๏ Assembly: within endoplasmic reticulum. Notable characteristics ๏ All viruses are serologically related ๏ Under Arboviruses (Arthropod-borned viruses, eg., mosquitoes, ticks) FLAVIVIRUSES Dengue (breakbone fever) Mosquito borne: ๏ Aedes aegypti, and Aedes albopictus Characterized by fever, severe headache, muscle and joint pain, nausea and vomiting, eye pain and rash (petechiae) Severe form of the disease: ๏ dengue hemorrhagic fever Occur to individuals with passively acquired (maternal Ab) or preexisting nonneutralizing heterologous dengue Ab due to previous infection with a different serotype virus Key features: increase vascular permeability → increase levels of vasoactive cytokines FLAVIVIRUSES Pathogenesis: Dengue (breakbone fever) Note FLAVIVIRUSES Pathogenesis: Development of Dengue hemorrhagic fever FLAVIVIRUSES Laboratory Diagnosis: Dengue (breakbone fever) RT-PCR—rapid determination Serotyping— in acute-phase serum, during period of fever Serological diagnosis ๏ Complicated due to cross-reactivity of IgG Ab to heterologous flavivirus Ag ๏ IgG ELISA and HI ๏ Dengue DOT (IgG/IgM) ๏ Dengue NS1 FLAVIVIRUSES Laboratory Diagnosis: Dengue (breakbone fever) FLAVIVIRUSES Overview: Hepatitis C virus Positive strand ssRNA, genus Hepacivirus Non-A, non-B hepatitis virus (NANB) Acquired through blood contact— blood transfusion (posttransfusion hepatitis), accidental needle prick Infections with varying characteristics: ๏ 75-85% will remain infected indefinitely ๏ Possible to have severe symptoms without permanent liver damage ๏ More common to have chronic liver disease, without overt symptoms ๏ Hepatocellular carcinoma may result from chronic HCV infection Interferon and Ribavirin— treatment to lessen liver damage No available vaccine FLAVIVIRUSES Pathogenesis: Hepatitis C virus FLAVIVIRUSES Serology: Hepatitis C virus Note FLAVIVIRUSES Laboratory Diagnosis: Hepatitis C virus Initial presentation ⇥ at 4 weeks ⇥ at 12 weeks post exposure ๏ HCV Ab, HCV RNA, and ALT CDC laboratory criteria for diagnosis: ๏ Anti-HCV ++ ๏ Hepatitis C virus detection test Nucleic acid test (NAT) for HCV RNA positive (including qualitative, quantitative, and genotype testing) Positive test indicating presence of hepatitis C viral antigen(s) This Slide is Intentionally Left Blank ! CORONAVIRUSES Introduction Properties ๏ Virion: spherical, helical nucleocapsid ๏ Genome: ssRNA linear, (+)sense, linear-nonsegmented ๏ Envelope: contains large, widely spaced, club- or petal-shaped spikes ๏ Replication: cytoplasm, budding into endoplasmic reticulum and Golgi ๏ Assembly: within endoplasmic reticulum. Notable characteristics ๏ Cause colds and SARS, MERS-Cov ๏ Difficult to grow in cell culture CORONAVIRUSES Illustration Human coronavirus ๏ Note the characteristic large, widely spaced spikes that form a “Corona” around the virion. credit: CDC - Dr. Fred Murphy CORONAVIRUSES Pathogenesis and Clinical Findings: SARS Highly species-specific ๏ Tropism for epithelial cells of the RT[HUMANS] or GIT Clinical findings ๏ Produce colds usually afebrile in adults. Symptoms similar to rhinoviruses —typified by nasal discharge and malaise ๏ Incubation period: 3-5 days, symptoms last ~1 week ๏ Causes severe respiratory disease Cases progress rapidly to acute respiratory distress— death (10%) CORONAVIRUSES Laboratory Diagnosis Molecular techniques ๏ PCR ๏ NAT Isolation and Viral ID ๏ Use of Vero monkey kidney cells Serology ๏ Testing on acute and convalescent sera— practical means of confirming coronavirus infections ๏ ELISA and Ht in which RC coated with CoV Ag are agglutinated by Ab- containing sera. This Slide is Intentionally Left Blank ! RETROVIRUS Introduction Properties ๏ Virion: spherical, cylindric core ๏ Genome: ssRNA linear, (+)sense ๏ Proteins: enveloped glycoprotein with Ag variation; Reverse-Transcriptase (RT) enzyme contained inside virions; protease required for viral production ๏ Envelope: present ๏ Replication: RT makes DNA copy from genomic RNA; provirus DNA is template for viral RNA with genetic variability ๏ Maturation: particles bud from plasma membrane RETROVIRUS Introduction Notable characteristics ๏ Members are nononcogenic and may be cytocidal ๏ Infect cells of the immune system (CD4+ containing cells) ๏ Proviruses remain permanently associated with cells ๏ Viral expression is restricted in some cells in vivo ๏ Cause slowly progressive, chronic diseases ๏ Replication is usually species-specific ๏ Group includes the causative agents of AIDS RETROVIRUS Illustration A HIV-infected lymphocytes ๏ Showing a large accumulation of freshly produced virus at the cell surface (A: 46,450x, bar=100nm); ๏ Newly formed virus budding from cytoplasmic membrane (B: 49,000x, bar= 100nm); B C ๏ Two virions about to be cast off from cell surface (C: 75,140x, bar=100nm) RETROVIRUS HIV genome and virion structure Synthesis of viral proteins ๏ Gag-Pol gene (ie., p11, p66, etc.) ๏ Gag gene (ie., p24, p17, etc.) ๏ Env gene (ie., gp120, gp41) Cleavage ๏ Gag-Pol & Gag: viral protease to produce small proteins (ie., p24) ๏ Env: cellular protease to produce SU gp120 and TM gp41 RETROVIRUS HIV cycle Binding Fusion RT Integration Transcription Translation Assembly Release RETROVIRUS Pathogenesis and Pathology Primary Infection Viral Dissemination Clinical Latency Opportunistic ⇥ (AIDS) Death RETROVIRUS Pathology Primary effects of HIV infection ๏ Extreme leukopenia— lymphocytes in particular ๏ Formation of giant T cells and other syncytial allowing the virus to spread directly from cell to cell ๏ Infected macrophages release the virus in CNS, with toxic effects, inflammation Secondary effects of HIV ๏ Destruction of CD4 lymphocytes allows for opportunistic infections and malignancies RETROVIRUS Pathology Signs and symptoms of HIV infection ๏ Symptoms are directly related to viral load level and of CD4 counts ๏ Initial infection— mononucleosis-like symptoms that soon disappear ๏ Asymptomatic phase 2-15 years (ave. 10) ๏ HIV destroys the immune system ๏ When T4 cells levels fall below 200uL (CD4 count), AIDS symptoms appear including fever, swollen lymph nodes, diarrhoea, weight loss, neurological symptoms (ie., Cryptococcal meningitis), opportunistic infections (eg., Pneumocystis carinii pneumoniae), and cancers (eg., Burkitt’s lymphoma) RETROVIRUS Serology Primary Infection Eclipse period Days Fiebig stage Months Clinical signs Symptoms Chronic phase Years AIDS RETROVIRUS Major gene products of HIV that are useful in diagnosis ๏ a— refers to the approximate molecular mass of the protein ๏ b— Ab to these proteins are the most commonly detected ๏ c— two-letter abbreviation of viral protein RETROVIRUS Laboratory Diagnosis Molecular methods ๏ NAT, RT-PCR Serology ๏ Anti-HIV (1&2) later flow IA (immunochromatographic) or ECLIA (Electrochemiluminescence Immunoassay) —3rd Gen. ๏ Vidas HIV DUO [Anti-HIV 1/2 + p24] — ELFA (Enzyme-Linked Fluorescence Assay) — 4th Gen. ๏ Western Blot— gold standard Algorithm est. by CDC and APHL in 80s remains unchanged 20years More is known about the disease and technology is evolving (increase sensitivity of the assay) Western Blot and IFA now less sensitive than some screening assays which they are intended to “confirm” RETROVIRUS Laboratory Diagnosis Molecular methods ๏ NAT, RT-PCR Serology ๏ Anti-HIV (1&2) later flow IA (immunochromatographic) or ECLIA (Electrochemiluminescence Immunoassay) —3rd Gen. ๏ Vidas HIV DUO [Anti-HIV 1/2 + p24] — ELFA (Enzyme-Linked Fluorescence Assay) — 4th Gen. ๏ Western Blot— gold standard RETROVIRUS Laboratory Diagnosis Confirmatory testing ๏ Western Blot ๏ Biorad Geenius™ HIV 1/2 Supplemental assay—used as a differentiation assay Resolution of “indeterminates” Ability to confirm HIV-2 infection Increase detection of acute infection Use of assays as screening or confirmatory / supplemental tests and as part of multi-test algorithms — rHIVda (Rapid HIV Diagnostic Algorithm ๏ Rapid Multiplex Test Alere Determine™ HIV 1/2 Ag/Ab Combo— rapid test that detects both HIV-1/2 Ab and the HIV-1 Ag (not intended for use in blood donor screening) RETROVIRUS Prevention: HIV infection NO vaccine available ๏ Monogamous sexual relations (ie., MSM) ๏ Condoms ๏ Universal precautions— health care workers No cure; therapies slow down the progress of the disease or diminish the symptoms Inhibit viral enzymes; RT, Protease INH, or Integrase INH Inhibit fusion Inhibit viral integration (Integrase INH) Highly active Anti-Retroviral therapy This Slide is Intentionally Left Blank ! Nonenveloped Nonsegmented ssRNA Introduction Picornaviruses ๏ Enteroviruses — poliovirus, HAV ๏ Rhinoviruses — rhinovirus ๏ Cardiovirus — infects heart and brain NE-NS ssRNA: Hepatitis A Virus Overview Cubical picornavirus relatively resistant to heat and acid Not carried chronically, principal reservoirs are asymptomatic, short-term carriers or people with clinical disease Self-limiting Fecal-oral transmission, multiplies in small intestine and enters the blood and is carried to the liver Most infections subclinical or vague Flu-Like symptoms occur Jaundice is seldom present No specific treatment once the symptoms begin Inactivated viral vaccine Attenuated viral vaccine Pooled immune serum globulin for those entering into endemic areas Reoviruses Overview Unusual, only dsRNA genome Two best known: ๏ Rotavirus Oral-fecal transmission Primary viral cause of mortality and morbidity resulting from diarrhoea in infants and children Treatment with dehydration and electrolyte replacement ๏ Reovirus Cold-like upper RT infection Enteritis Prions and Spongiform Encephalopathies Overview Prions ๏ Proteinaceous infectious particles, highly resistant to chemicals, radiation, and heat. ๏ Cause Transmissible Spongiform Encephalopathies (TSE) in humans and animals Neurodegenerative disease with long incubation periods Creutzfeldt-Jakob disease (CJD)— alteration in the structure of normal PrP protein found in the brain ‣ Abnormal PrP converts normal PrP into abnormal form Abnormal PrP results in nerve cell death, spongiform damage, and severe loss of brain function Transmission in through direct or indirect contact with infected brain tissue or CSF Prions and Spongiform Encephalopathies Overview CJD ๏ Variant CJD became apparent in the late 1990s after eating meat from cattle afflicted with bovine spongiform encephalopathy ๏ Difficult to diagnose, requires examination of brain or nervous tissue biopsies ๏ Prevention relies on avoidance of contaminated tissue ๏ Treatment focuses on easing symptoms Thank You! ⇥ Fin~