Virology - HO3-VIROLOGY (FINALS) PDF

VIRUSES the smallest infectious agents and contain only one type of nucleic acid (DNA or RNA). Recognition of their presence is only possible either by Direct Electron Microscope obligate intracellular parasite multiply only in living cells FUNDAMENTALS OF VIROLOGY A. VIRION a mature virus parti...

VIRUSES the smallest infectious agents and contain only one type of nucleic acid (DNA or RNA). Recognition of their presence is only possible either by Direct Electron Microscope obligate intracellular parasite multiply only in living cells FUNDAMENTALS OF VIROLOGY A. VIRION a mature virus particle containing a nucleic acid core surrounded by a protein coat, with or without an envelope NUCLEIC ACID Each virus contains only one kind of nucleic acid (DNA or RNA), containing the entire viral genome; nucleic acid in viruses are classified as either: SINGLE STRANDED VS. DOUBLE LINEAR VS. CIRCULAR SEGMENTED VS. NON-SEGMENTED NUCLEOCAPSID Each nucleocapsid contains a nucleic acid core surrounded by a protein coat, the capsid (which consists of many subunits named capsomeres); the; capsid symmetry is either. HELICAL ICOSAHEDRAL COMPLEX ENVELOPE lipid-containing membrane that surrounds some virus particles. It is acquired during viral maturation by a budding process through a cellular membrane. Some viruses have envelopes which can ether be from the: Host cell membrane Host nuclear membrane Some viruses have no envelope - "naked" NAKED VIRUSES ENVELOPED VIRUSES Environmentally stable to the following: Temperature Gastric acidity Proteases Detergents Drying Environmentally labile Modifies cell membrane during replication; released by budding and occasionally, cell lysis Destroy cell by lysis Can easily spread by fomites, hand-to-hand contact, dust and small droplets; can dry out and retain infectivity VIRAL SIZE Can be visualized directly by electron microscopy Size ranges from 20-300 nm POX VIRUS: are the largest DNA viruses PARAMYXO: are the largest RNA viruses PARVO: are the smallest animal virus B. TAXONOMY TAXONOMIC LEVEL SUFFIX EXAMPLE Order virales Mononegavirales Family viridae Paramyxoviridae Subfamily vinnae Paramyxovirinae Genus virus Morbillivirus Species Based on disease produced or derived from where they were found or after a particular name Measles virus West Nile Virus Coxsackie virus Norwalk Virus Epstein-Barr virus C. REPLICATION CYCLE OF VIRUSES Viruses are obligate intracellular parasites; they require other cells to reproduce since they do not have their own metabolic machineries ABSORPTION interaction of a virion with a specific receptor site on the surface of the cell. Receptors are generally glycoproteins Receptors used by some viruses CMV Integrins (heparan sulfate) EBV CD21 HIV CD4, CXCR4, CCR5 Parvovirus B19 P antigen on RBCs Rabies Nicotinic AChR Rhinovirus ICAM-1 PENETRATION aka engulfment virus particles is taken up inside the cell UNCOATING physical separation of the viral nucleic acid from the outer structural components of the virion. So that it can function SYNTHESIS Early mRNA and protein synthesis Viral genome replication Late mRNA and protein synthesis ASSEMBLY OF VIRION RELEASE either through host cell lysis (usually for naked viruses) or through budding (for enveloped viruses) LABORATORY DIAGNOSIS OF VIRAL INFECTIONS Clinical Symptoms For Virus Isolation Direct Examination For Serology Viruses Commony Assoc. Paired sera Influenza Parainfluenza RS virus* Adenovirus Rhinovirus Paired sera Rotavirus Norwalk agent Adenovirus Enterovirus Varicella-zoster Herpes simplex Measles Rubella or Enterovirus Throat swabs, nasal washings, nasopharyngeal aspirate (babies <2 years old) Throat washing (EM) nasopharyngeal aspirate (IF) Stool Rectal swabs Throat swabs Stool (EM) (ELISA) Skin lesions or rash Skin scrapings, Vesicular fluid or swabs Rectal swabs (enterovirus only) Vesicular fluid (EM) Ulcer scraping (EM) (IF) Paired sera Encephalitis or aseptic meningitis Brain biopsy Blood & CSF (togavirus) Brain biopsy (EM) Paired sera Throat and rectal swabs Corneal impression smears (IF for babies) Stool Serum for HBsAg (EM or RIA) Stool for Hepatitis A (EM) Throat swabs, Products of conception None Urine Urine (EM for CMV) Respiratory syndromes Gastroentiritis Hepatitis Congenital infection SPECIMEN COLLECTION, STORAGE AND TRANSPORT Herpes simplex virus Togavirus Enterovirus Rabies virus Serum Hepatitis A Hepatitis B Single serum (mother & infant) Rubella Cytomegalovirus Viruses are fastidious; sensitive to heat and drying Clinical specimens should be handled with care and delivered to the laboratory as soon as possible 4°C is satisfactory for a few hours -70°C is necessary for days of transport Throat swab or washings -keep in sterile saline containing antibiotics and protein stabilizer Rectal swabs (stool samples preferable) in sterile saline) Skin scrapings or lesion swabs; place in sterile tubes with protein stabilizers Urine - collect in sterile containers 7 Body fluid or vesicle - collect in sterile containers Blood - heparinized blood for plasma; clotted blood for serum Others - biopsy tissues - keep in sterile saline autopsy tissues - keep in sterile containers without preservatives CELL CULTURE SELECTION FOR VIRUS ISOLATION AND PRESUMPTIVE DIAGNOSIS Many viruses can be grown in cell cultures or in fertile eggs under strictly controlled conditions. Growth of virus in is animals is still used for the primary isolation of certain viruses and for studies of the pathogenesis of viral diseases and of viral oncogenesis. EXAMPLES: Monkey Kidney Cells Human Embryo Fibroblasts Hep-2 line - originally consisted of laryngeal epidermal carcinoma cells A549 line - originated from type Il alveolar adenocarcinoma cells Rabbit Kidney Cells BASIC TYPES OF CELL CULTURES PRIMARY CLUTURE made by dispersing cells (usually with trypsin) from freshly removed host tissues. In general, they are unable to grow for more than a few passages in culture. DIPLOID CELL LINE secondary cultures that have undergone a change that allows their limited culture (up to 50 passages) but that retain their normal chromosome pattern. DIPLOID CELL LINE secondary cultures that have undergone a change that allows their limited culture (up to 50 passages) but that retain their normal chromosome pattern. CELL CULTURE SELECTION No single cell culture system is susceptible to all viruses or even most virus types infecting humans Different cell culture types must be used when an unknown virus is suspected Different virus types have different host cell spectrums for their replication Selection of appropriate cell culture systems essential CYTOPATHIC EFFECTS (CPE) morphologic changes in infected cells; types of virus-induced cytopathic effects include: Cell lysis or necrosis Inclusion formation Giant cell formation e.g. observed in HSV infection Cytoplasmic vacuolization EXAMPLE OF INCLUSION BODIES Intracytoplasmic inclusion bodies Intranuclear inclusion bodies Both intranuclear and intracyloplasmic Negri bodies Rabies Virus Molluscum bodies Molluscum contagiosum virus Guarneri bodies Vaccinia virus Owl's eye Cytomegalovirus Cowdry type A bodies Herpes simplex virus Varicella-Zoster virus Cowdry type B bodies Polio Virus Torres Bodies Yellow fever virus Warthin-Finkeldey Bodies Measles Virus METHODS OF DIAGNOSIS morphologic changes in infected cells; types of virus-induced cytopathic effects include: Cell lysis or necrosis Inclusion formation Giant cell formation e.g. observed in HSV infection Cytoplasmic vacuolization 1. CYTOLOGY/HISTOLOGY Cytology: HSV. VZV Histology: adenovirus, CMV, measles, papilloma virus, parvovirus, rabies Microscopic examination for viral inclusions. Less sensitive than culture but helpful for viruses that are difficult to grow 2. ELECTRON MICROSCOPY Gastroententis: calicivirus, coronavirus, enteric adenovirus, Norwalk agent Encephalitis: JC polyoma virus, HSV, measles Little used, labor-intensive and expensive Most useful for viruses that can be grown 3. IMMUNOFLUORESCENT STAINS Adenovirus, CMV, HSV 1 and 2, influenza A and B, parainfluenza, RSV. VZV Antiviral antibodies can be used to stain viral antigens in specimens and/or cultures 4. ENZYME IMMUNOASSAY Enteric adenoviruses, HSV, rotavirus, RSV Solid phase and membrane ELISAs are most common 5. ENZYME-LINKED VIRUS INDUCIBLE SYSTEM HSV 1 and 2 Baby hamster kidney cell culture with betagalactosidase gene that is only expressed when cells are infected with virus 6. CONVENTIONAL CELL CULTURE Most common: adenovirus, CMV, enteroviruses, HSV, Influenza A and B, parainfluenza, RSV 1-28 days for cytopathic effects: cell rounding, clumping, vacuolation, granulation, giant multinucleated cells, cell fusion, syncytial formation, cell lysis; plaques, Inclusion bodies 7. SHELL VIAL CULTURE Any that grow via conventional cell culture CMV and VZV Rapid, within 1-2 days 8. LATEX AGGLUTINATION Rotavirus Direct test, latex coated with antibodies, rapid and inexpensive 9. POLYMERASE CHAIN REACTION Enterovirus in CSF Limited commercial availability 10. VIRAL SEROLOGY Arbovirus, CMV, EBV, Hepa, HSV, measies, rubella, V2V (Detects in serum. Useful in evaluating immune status or diagnosing viral infections where culture is difficult or impossible. DNA VIRUSES 1. ADENOVIRIDAE 70-90 nm Cubic symmetry Viruses: Adenoviruses serotype 1-48 Infects the Respiratory tract and urinary tract 2. HEPADNAVIRIDAE 40-48 nm Cubic symmetry Hepatitis B virus Hepatitis B 3. HERPESVIRIDAE 150-200 nm Cubic symmetry HSV 1 & HSV 2: VARICELLA ZOSTER VIRUS EPSTEIN-BARR VIRUS: CYTOMEGALOVIRUS Human Herpes Virus 6,7,8 4. PAPOVAVIRIDAE 45-55 nm Human papilloma virus (HPV) Polyomavirus (BK and JC viruses infect humans) Genital warts, anogenital wars, skin warts Hemorrhagic cystitis (BKV) or progressive multifocal eukoencephalopathy (JCV) 5. PARVOVIRDAE 18-26 nm Cubic symmetry Parvovirus B-19 Erythema infectiosum (fifth disease) 6. POXVINDAE 220-450 nm long x 140-260 nm wide x 140-260 nm thick Complex structure Variola Major:Smallpox Variola Minor: Alastrim (mild form of smallpox) Vaccinia: Cowpox RNA VIRUSES 1. ARENAVIRUS 50-300 nm Lymphocytic choriomeningitis (LCM) Lassa fever viruses Hemorrhagic fever viruses Aseptic meningitis, influenza-like symptoms Lassa hemorrhagic fever Bolivian and Argentinian hemorrhagic fever 2. BUNYAVINDAE 90-120 nm Califomia encephalitis virus (Lacrosse Virus): Encephalitis Group C Arbovirus Hanta Virus: Hanta Virus Pulmonary Syndrome 3. CALICIVIRIDAE 35-39 nm Norwalk Agent: Gastroententis Hepatitis E Virus: Hepatitis 4. CORONAVINDAE 80-130 nm Pleomorphic with large club shaped projections Coronavirus Coryza: Severe Acute Respiratory Syndrome (SARS) 5. FILOVIRIDAE 45-55 nm Spherical Group B Arbovirus Yellow fever virus: Yellow Fever Dengue fever virus: Dengue Fever St. Louis encephalitis virus: Meningitis, Encephalitis, Encephalomyelitis West Nile virus (WNV): Generally asymptomatic fever, muscle pains Hepatitis C Virus: Hepatitis 6. ORTHOMYXOVIRIDAE 100 nm Spherical/pleomorphic Influenza A: Influenza (malaise, headache, myalgia, cough) Influenza B: Similar to "mild' influenza 7. PARAMYXOVIRIDAE 150-300 nm Spherical/pleomorphic: Measles virus: Measles; pneumonia, encephalitis, subacute sclerosing panencephalitis RSV (Pneumovirus): URTI and LRTI especially in infants Mumps virus (Rubulavirus): Mumps; Orchitis, oophoritis, meningitis, deafness Parainfluenza virus 1-4: Common cold, tracheobronchitis, rarely pneumonia 8. PICORNAVIRIDAE 20-33 nm Poliovirus: Polio Coxsackievirus A: Herpangina, hand-fool-and-mouth disease Coxsackievirus B: Pleurodynia (Bornholm disease, epidemic myalgia) Foot-and-mouth disease virus: Foot-and-mouth disease (rarely in humans) Hepatitis A Virus: Acute hepatitis and Jaundice Rhinovirus: Coryza 9. REOVIRIDAE 60-80 nm Variable morphology, spokes wheel Reovirus: Little association with disease Rotavirus: Gastroenteritis (infants and children 6mos-2yrs) Orbivirus: Colorado Tick Fever 10. RETROVIRIDAE 80-100 nm Slightly pleomorphic to spherical HTLV-1: Adull T-cell leukemia (Sezary 7 cell leukemia), lymphoma HTLV-2: Hairy Cell leukemia HIV-1: AIDS Stage 1: Asymptomatic Stage 2: AIDS-Related Complex Stage 3: Full Blown AIDS HIV-2: Immunodeficiency (less pathogenic than HIV-1) 11. RHABDOVIRIDAE 50-95 nm x 130-389 nm Bullet shaped Rabies virus: Rabies 12. TOGAVIRIDAE 60-70 nm Spherical Alphavirus (Group A Arbovirus) Eastern equine encephalitis Western equine encephalitis, Venezuelan equine encephalitis Rubella Virus (Rubivirus) German Measles