L01 Ch23 Serological and Molecular Detection of Viral Infections PDF

6/27/2024 SEROLOGICAL AND MOLECULAR DETECTION OF VIRAL INFECTIONS Chapter 23 PowerPoints are a general overview and are provided to help students take notes over the video lecture ONLY. PowerP...

6/27/2024 SEROLOGICAL AND MOLECULAR DETECTION OF VIRAL INFECTIONS Chapter 23 PowerPoints are a general overview and are provided to help students take notes over the video lecture ONLY. PowerPo int s DO NOT c over the details nee de d fo r t he Un it e xam Each student is responsible for READING Preamble the TEXTBOOK for details to answer the UNIT OBJECTIVES Unit Objectives are your study guide (not this PowerPoint) Test questions cover the details of UNIT OBJECTIVES found only in your Textbook! 1  6/27/2024 Virus structure and life cycle Immune defenses against viruses Viral escape mechanisms Chapter Laboratory testing for viral infections Hepatitis viruses Overview Hepatitis A, E, B, D, C Herpes viruses EBV, CMV, VZV Other viral infections Rubella, rubeola, mumps, HTLV-I/II Virus Structure Submicroscopic particles(nm) Core of DNA or RNA Protein coat (capsid) Some have outer envelope 2  6/27/2024 Virus Life Cycle Obligate intracellular pathogens Rely on host cells for their replication and survival Exist as freely circulating particles and intracellular particles Innate defenses Skin and mucous membrane barriers Recognition of PAMPs on virus-infected host cells Immune Interferons α and β Defenses Humoral antibody responses Antibodies attack free virus particles Against Viral neutralization, opsonization, C’ fixation, ADCC Viruses Cell-mediated immunity Interferon γ and IL-2 produced by Th1 cells Host cells containing intracellular virus are destroyed by CTLs 3  6/27/2024 SUMMARY OF IMMUNE DEFENSES AGAINST VIRUSES Viral Escape Mechanisms 1. Mutations result in production of new viral antigens. e.g., Inf luenza viruses undergo frequent genetic changes. 2. Viruses block action of immune system components. e.g., HSV can bind C3b. 3. Suppression of the immune response e.g., CMV reduces MHC I expression. 4. Immune function altered e.g., EBV stimulates polyclonal B-cell activation. 5. A latent state is established. e.g., VZV remains latent in nerve cells. 4  6/27/2024 Serological tests Distinguish between current and past infection IgM (+), IgG (+/–): current or recent infection; congenital infection Laboratory IgM (–), IgG (+): past infection Antibody titers used to monitor course of infection Testing for Assess immune status Virus-specific IgG indicates immunity Viral Molecular tests Infections Detect active infection Quantitative tests—guide antiviral therapy Hepatitis Viruses Hepatitis in the inflammation of the liver; caused by: – Radiat ion * Hypot h ermia – C h emical t oxins * Bact eria – Secon dar y to ot her disease * Fun gi – C irrhosis ( alc oho lism) * Parasit es – Drugs 5  6/27/2024 Hepatitis Viruses Transmitted by fecal-oral Transmitted via parenteral route route Hepatitis A virus (HAV) Hepatitis B virus (HBV) Hepatitis E virus (HEV) Hepatitis D virus (HDV) Hepatitis C virus (HCV) General flu-like symptoms early in infection Clinical and Pain in upper right quadrant of abdomen Laboratory Hepatomegaly and liver tenderness with progression Findings of Jaundice, dark urine, light feces Hepatitis Elevated bilirubin and liver enzymes (e.g., ALT) 6  6/27/2024 Hepatitis A: HAV RNA virus transmitted by: Fecal-oral route Close person-to-person contact Ingestion of contaminated food or water Produces acute hepatitis in majority of adults. Infections in children are usually asymptomatic. Formalin-inactivated HAV vaccine Hepatitis A: HAV (continued) HAV immune globulin may be recommended for unimmunized persons exposed to the virus. Acute infection is indicated by (+) IgM anti-HAV. Immunity is indicated by (+) total anti-HAV along with (–) IgM anti-HAV. 7  6/27/2024 RNA virus with four genotypes HEV-1 and HEV-2 are transmitted primarily through ingestion of feces-contaminated drinking Hepatitis E: water. HEV-3 and HEV-4 are transmitted mainly by consumption of infected pork. HEV Mostly asymptomatic or self-limiting infections Severe infections possible Pre gnant women Hepatitis E: Immunocompromised persons Acute infection is indicated by IgM anti-HEV. HEV IgG anti-HEV indicates past exposure. (continued) Can detect HEV RNA in blood or stool samples during acute infection. 8  6/27/2024 DNA virus transmitted through parenteral or perinatal routes Sexual contact Hepatitis B: IV drug use HBV Occupationa l nee dlestick injur y During bir th process Acute infection: symptoms increase with age Chronic infection Persists for 6 months or more Hepatitis B: HBV Occurs in 90% of infected infants, 10% of infected adults Increases risk of liver cirrhosis or hepatocellular carcinoma (continued) Infection preventable by immunization Hepatitis B immune globulin (HBIG) may be given to unimmunized persons exposed to HBV. 9  6/27/2024 Hepatitis B surface antigen (HBsAg) First HBV marker to appear Serological Protein on outer envelope of virus Excess circulates in virus-like particles in blood Markers for Marker for active HBV infection Component of hepatitis B vaccine HBV Hepatitis Be antigen (HBeg) Antigens Protein in the core of the HBV Marker of active viral replication Indicates high degree of infectivity Anti-HBc Directed against hepatitis B core antigen IgM anti-HBc indicates current or recent acute infection; “Core window” - period when neither hepatitis B surface antigen (HBsAg) nor its antibody (HBsAb) can be detected in the serum of the patient only the Antibody Anti-HBc Total anti-HBc consists mainly of IgG and can indicate a current or a past infection. Markers for Anti-HBe HBV Directed against HBeAg Indicates recovery from hepatitis B. Anti-HBs Directed against HBsAg Indicates immunity to hepatitis B. 10  6/27/2024 TYPICAL MARKERS IN ACUTE HBV INFECTION TYPICAL MARKERS IN CHRONIC HBV INFECTION 11  6/27/2024 CHEMILUMINESCENT IMMUNOASSAY FOR HBSAG Hepatitis D: HDV RNA virus that requires presence of HBV Transmitted through parenteral or perinatal routes Coinfection with HBV U su ally results in ac ut e, self- limited hepat itis ( +) for an ti- HDV and IgM ant i- HBc Superinfection of chronic HBV carriers C h ro nic liver disease wit h acc elerat ed pro gressio n t o c irrh osis and liver failure ( +) for an ti- HDV and IgG an ti- HBc HDV RNA = marker of active viral replication 12  6/27/2024 Hepatitis C: HCV RNA virus with seven genotypes Transmitted by exposure to contaminated blood, sexual contact, perinatal exposure Most infections are asymptomatic at first but develop into chronic liver disease. Anti-HCV IgG is used for screening and diagnosis. Hepatitis C: HCV (continued) Qualitative HCV RNA is used for confirmation. Quantitative molecular tests are used to monitor viral load during antiviral therapy. Genotyping is used to determine best therapy. 13  6/27/2024 Epstein-Barr Virus (EBV) DNA herpes virus most commonly transmitted by intimate contact with salivary secretions Begins in oropharynx in B cells and epithelial cells and spreads through lymphoreticular system. Causes a spectrum of diseases Infectious mononucleosis Lymphoproliferative disorders Cer tain malig nancies (e.g., Burkitt lymphoma) Infectious Mononucleosis (IM) Classic sympto ms – Fever – Lymphadenop athy – Sore throat – Fatig ue Labo rat or y f indin gs – Absolute lymphocytosis – 20% or more atypical lymphocytes – Heterophile a ntib ody  Reacts with antigens from two or more species  Monospot  Paul-Bunnell test 14  6/27/2024 Antibodies to EBV Antigens E a r l y a nt i g e ns EA-D (early antigen diffuse) – A n t i - E A -D i s p r es e n t i n a c u t e IM. EA-R (early antigen restricted) L a t e a n t i g e ns e. g., v i r a l c a p s i d a n t i g e n s ( V C A ) Ig M a n t i - V C A i s b e s t m a r k e r f o r a c u t e I M. Ig G a n t i - V C A c a n i n d i c a t e a c u t e o r p a s t I M. L a t e nt - p ha s e a nt i g en s – e. g., E B V n u c l ea r a n t i gen s ( E B N A) – A n t i - E B NA a p p ea r s d u r i n g c o nv a l es c en t IM Cytomegalovirus (CMV) D N A h er p es vi r us i s t ra n s m i t t ed t h ro ug h o ra l s e c r e t i o ns , g eni t a l s ec r e t i o ns , c o ng en i t a l l y, o r b y t ra ns f us i o n / t ra ns p l a nt a t i o n. H e a l t hy i n d i vi d u a l s m a y b e a s y m p t o m a t i c o r d e v e l o p a m o no nuc l e o s i s - l i ke s y n d r o m e. In i m m uno s up p r e s s e d p er s o ns ( e. g., t ra ns p l a nt p a t i ent s , p a t i ent s w i t h A I D S ) , C M V c a n d i s s e m i na t e t o l un g s , l i v er, G I t ra c t , C N S, a nd e ye s a nd c a us e l i fe - t hr e a t eni ng i nfe c t i o ns. C M V m a y c a u s e c o ng eni t a l d efec t s a nd d e c re a s e d s ur v i va l i n infants. 15  6/27/2024 Direct virus detection Vira l cu ltu re Sample plac ed wit h a c ell t ype th at t h e viru s being Laboratory t est ed for can in fec t. If th e cells show changes, kn own as cyt opat hic Diagnosis effect s, t hen th e cu ltu r e is positive. Iden tif ic at ion of CMV a n tigens of CMV Molecu lar t est s for CMV DN A Serology U sed t o sc reen blood a nd organ don ors; pregna nt women P resence of IgG an ti- CMV indicat es in fect ion. Low a v idit y an tibodies indica te rec en t infect ion. Varicella-Zoster Virus (VZV) D N A her p es vi r us Tra ns m i t t e d b y i nha l a t i o n o f i nfe c t ed r es p i ra t o r y s ec r e t i o ns o r a er o s o l s f r o m s k i n l e s i o ns C a us e o f : Va r i ce l l a ( ch i cken p ox ) He rp es z os t e r (s h i n g l es ) P r e ve nt a b l e b y va c c i na t i o n 16  6/27/2024 Laboratory Diagnosis of VZV Diagnosis is usually based on characteristic clinical findings. Laboratory method of choice is qPCR to detect VZV DNA in clinical samples. Serology is most useful in determining immunity to VZV in health- care workers, pregnant women, and organ transplant candidates. RNA virus transmitted through respiratory droplets or across the placenta Cause of German measles Can cause: Rubella Congenital abnormalities Deafness, eye defects Cardiac abnormalities Virus Mental retardation Motor disabilities Miscarriage Stillbirth in infants born to infected mothers Infection preventable by immunization 17  6/27/2024 Serology Presence of IgG is used to screen for immunity. Laboratory Congenital infection is indicated by IgM antibodies in fetal blood, cord blood, or neonatal serum. Current infection is indicated by rubella-specific IgM or fourfold rise Testing for in IgG. Low avidity antibodies indicate recent infection. Rubella Viral culture Molecular methods Rubeola RNA virus transmitted through respiratory droplets Cause of:  Mea s l es  S u ba c u t e s cl er os i n g p a n e n ce p h al i t i s ( SS P E ) Infection preventable by immunization Diagnosis is usually based on clinical presentation and confirmed by serology. 18  6/27/2024 Mumps RNA virus transmitted through respiratory droplets, saliva, and fomites Most common clinical manifestation is parotitis. Infection is preventable by immunization. Diagnosis is usually based on clinical presentation. Confirmation is done by culture or RT-PCR. HTLV-I/II Closely related retroviruses that preferentially infect T lymphocytes Transmission is mainly bloodborne, via sexual contact, or perinatal (especially breastfeeding). HTLV-I is the cause of adult T-cell leukemia /lymphoma (ATL) and HAM/TSP. Serological tests for antibodies to HTLV-I/II are used to diagnose infections and screen blood donors. ELISA or CLIA are used to screen; Western blot or LIA are used for confirmation of positive results. 19  6/27/2024 Toxoplasmosis Found in feces of h o u s e ca t s (d ef i n i t i v e h o s t ) r o de n t s other mammals Mode of infection: A cci d en t a l i n g es t i o n o f o o cy s t (f ec a l co n t a mi n a t i o n o f m ea t , raw mi l k , et c.) Tra n s pl a c en t a l t ra n s mi s s i o n Toxoplasmosis Prevention of congenital toxoplasmosis avoid touching mucous membranes of mouth and eyes while handling raw meat wash hands and kitchen surfaces af ter coming into contact with raw meat be careful not to allow flies and cockroaches access to food avoid contact (or wear gloves) when handling cat litter boxes or gardening Signs and symptoms usually asymptomatic, organism can multiply in any organ of body if symptoms, similar to mono spontaneously recovery 20  6/27/2024 Toxoplasmosis Congenital infection - of most concern r es u l t i n C NS ma l f o r ma t i o n a n d mo r t a l i t y o f t h e n eo n a t e 75 % h av e n o s y mp t o ms a t b i r t h , d i s ea s e i s do r ma n t , d i s cov e re d o n l y w h e n n eu ro l o gi c a l p ro b l em s s u c h a s b l i n d n es s o ccu r s Laboratory diagnosis E IA TORCH Testing So m etim es ca lled t h e TORCH p a ne l Con s is ts o f t es t s for a nt ibod ie s to fo ur or ga nis m s tha t ca us e congenit a l infectio ns tra n s mit ted fr om mo ther t o fet us. Na me o f th e t es t is a n a cr onym for the o r ga nis ms detecte d by t his pa nel: Toxoplasma g o nd ii (toxopla s m os is ), r u bella (G er m a n m ea s les ), cyt om eg a lo vir us (CMV ), a nd he r pe s s imp le x vir us (HSV ). O s ta n ds fo r ot h er s wh ich u s u a lly include s s yphilis , hep a titis B, coxs a ckie vir us , Ep s tein -Ba r r vir u s , va r icella -z o s te r vir us , a nd hum a n pa r vovir us. 21  6/27/2024 Other Viruses 1. Adenovirus - respiratory 2. Arbovirus - skin - mosquitoes, ticks 3. Influenza - respiratory 4. Rotovirus - gastrointestinal (fecal /oral route) 5. Mumps - respiratory 6. Measles - respiratory 7. RSV - respiratory(Respiratory Syncytial Virus) Viruses are obligate intracellular pathogens that can cause many human diseases. Antibodies bind to free virions to prevent spread of infection, whereas cytotoxic T cells or NK cells are required to attack Summary intracellular viruses. Viruses have evolved several ways to escape host defenses, and some may establish a latent state in the host genome. 22  6/27/2024 Viral infections can be identified directly by culture, antigen detection, or molecular methods to detect viral nucleic acid. Summary Serological tests are often used to indicate exposure to viral pathogens. (continued_1) Presence of virus-specific IgM antibodies indicates a current or recent infection or a congenital infection. Presence of IgG antibodies indicates a previous exposure or immunity to a virus. Serology plays a key role in the diagnosis of viral hepatitis caused by the fecally transmitted HAV or HEV or the parenterally transmitted HBV, HDV, or HCV. Summary EBV is the cause of IM and various (continued_2) malignancies; serological tests for the heterophile antibody and EBV-specific antibodies play a key role in the diagnosis of IM. 23  6/27/2024 CMV is a herpes virus that can cause congenital infections and disseminated disease in immunocompromised persons. Summary CMV infection is best detected (continued_3) by molecular assays and tests for CMV antigens. Serology is useful in identifying individuals who have been exposed to the virus, such as potential organ donors. Many viral infections are preventable by routine immunization. Serology plays a key role in identifying persons who are immune to these infections: Varicella (chickenpox and shingles) Summary Rubella (German measles and congenital rubella) Rubeola (measles and SSPE) (continued_4) Mumps HTLV-I is the cause of adult T-cell leukemia and HAM/TSP, a progressive neurological disease. Serological tests are helpful for diagnosing these diseases and screening blood donors. 24  6/27/2024 READ the TEXTBOOK for the details to answer the UNIT OBJECTIVES. USE THE UNIT OBJECTIVES AS A STUDY GUIDE Postamble All test questions come from detailed material found in the TEXTBOOK (Not this PowerPoint) and relate back to the Unit Objectives 25