MDMICRO Medical Microbiology Long Exam 4 (RNA Viruses) PDF

MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY MDMCIRO LE4...

MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY MDMCIRO LE4 ○ With vectors @ August 8 (8:00-10:00AM - SJ504) 1) Family Arenaviridae - Lab diagnosis discussed a) Genus Arenavirus - immunity prevention for vaccines discussed Viruses: Chapre, Guanarito, Junin, Machupo, Sapia, and Whitewater Arroyo - treatment & control Many are rodent-borne NOT PART: **kung ano lang discuss na family and genus yung lalabas sa exam** 2) Family Bunyaviridae (4) a) Genus Orthobunyavirus - epidemi Known arthropods are Anopheles A and B - pathology Diseases: Encephalitis, Guama - prions b) Genus Hantavirus Rodent borne Family Flaviviridae …………………………………………………………………………………………………………………………….……..……2 c) Genus Nairovirus Family Retroviridae…………………………………………………………………………………………………………………………………...10 Artrophod borne carried by ticks causing Nairobi sheep disease and Family Orthomyxoviridae…………………………………………………………………………………………………………………………….13 hemorrhagic fever Family Picornaviridae………………………………………………………………………………………………………………………………………..15 d) Genus Phlebovirus Family Rhabdoviridae…………………………………………………………………………………………………………………………………….16 Arthropod borne: sandfly and ticks Coronaviridae……………………………………………………………………………………………………………………………………………………17 Diseases: Rift Valley fever virus, severe fever with thrombocytopenia (low platelet) syndrome virus 3) Family Filoviridae (2) – Filo ME Classification and Properties of Some Arthropod-Borne (Arboviruses) and Rodent-Borne a) Genus Marburgvirus Viruses b) Genus Ebolavirus 4) Family Flaviviridae [+SS RNA - Class IV] a) Genus Flavivirus Arthropod borne: mosquitoes and ticks Dengue, Zika viruses, Japanese B encephalitis 5) Family Reoviridae (2) OCay a) Genus Coltivirus Colorado tick fever virus Arthropod borne - ticks and mosquitoes b) Genus Orbivirus Arhtropod borne - mosquitoes 6) Family Togaviridae a) Genus Alphavirus Diseases are arthropod borne carried by mosquitoes Chikungunya, eastern, western, and venezuela encephalitis Notice common between families below: enveloped + icosahedral + (+) SSRNA Summary; Exceptions & Variations ○ Family Flaviviridae (children > adults) Arthropod-borne viruses or Arboviruses MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY > Dengue: Helical + Longest SSRNA + Spike protein for ACE2 + 3 vaccines + host Hemagglutinin (protein E) + Non- & Structural proteins + Protease (host protease + qRT PCR & viral NS2B/3 protease) + Virion-Endosomal Membrane fusion (endocytosis) + 3 phases (FCR) + RT PCR + viremia ✔ Envelope ✖ Naked Replication in intracellular vesicle 1. Family Flaviviridae > dengue, japanese B 1. Family Picornaviridae > poliomyelitis ○ Bite → replicate in skin/langerhan cells → macrophage → encephalitis, Zika bloodstream – pwede rin hepatocyte & endothelial cells 2. Family Retroviridae > HIV Haematogenous spread → PNS, ocassionally CNS 3. Family Orthomyxoviridae > influenza > Jpanese B encephalitis 4. Family Rhabdoviridae > rabies 5. Family Coronaviridae > coronavirus Encephalitis; Blood-brain barrier (CNS neurons) + Overwintering Mechanism (dormant) + infective pigs → bird → human + human (+) SSRNA (-) SSRNA | Class V w/ RNA-dep. RNA polymerase or Replicase dead-end host + vertical transmission + ELISA w/ CSF + booster ○ Bite → blood/macrophage → blood-brain barrier → brain 1. Family Flaviviridae 1. Family Orthomyxoviridae > influenza > Zika 2. Family Picornaviridae > poliomyelitis 2. Family Rhabdoviridae > rabies 3. Family Coronaviridae > coronavirus Sexual intercourse + Guillain-Barre Syndrome (autoimmune against NS) 4. Family Retroviridae > HIV + Transplacental + Microcephaly (progenital/neural cells) & congenital Class IV Class VI abnormalities + vertical transmission mRNA → protein mRNA → RT → protein ○ Family Retroviridae > HIV Helical nucleocapsid + Reverse Transcriptase + (transmembrane) spike 1. Family Flaviviridae 1. Family Retroviridae 2. Family Coronaviridae proteins → CD4 receptor + RT qPCR + 3 genes 3. Family Picornaviridae Provirus = dormant @chromosome Replication in antigen-presenting cells (macrophages & Th lymphocytes) (+) SSRNA x2 IMPORTANT: Common cold viruses ○ Family Orthomyxoviridae > influenza (1) Rhinoviruses (most common) Helical symmetry, round filamentous + sialidase → host-cell-sialic acid (2) Enteroviruses (NANA—) + Antigenic surface proteins (immunogenic hema. & neura. → mucin (3) Coronaviruses → epithelial), Antigenic variation; drift, shift (4) Adenoviruses Variable antigen = influenza A (most) & influenza B (sometimes) → epidemic Variable stable → influenza C (mild) (-) SSRNA w/ 7-8 linear segments = Genetic reassortment (2 virus:1 cell = Family Flaviviridae parental + progeny) ○ Family Picornaviridae > poliomyelitis Genus Flavivirus Many virus types + 2 vaccines → IgA + 2 antibodies (secretory & serum) + cell-mediated immunity + viremia ○ Family Rhabdoviridae > rabies: Bullet shape + Spike Glycoproteins + Negri body (nucleocapsid aggregates) + Targets muscles → PNS → NS → brain (stem, cerebellum etc.) ○ Family Coronaviridae > coronavirus MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY Dengue Virus Japanese B encephalitis virus Zika Virus each other will code for different proteins Vectors: Aedes Vectors: Culex Vector: Aedes aegypti 2) Translation of the viral genome → long polypeptide → then divided (proteolytic aegypti & Aedes tritaeniorhynchus & Aedes albopictus processing of viral polyprotein by host proteases and the viral NS2B/3 protease) to albopictus generate structural and nonstructural proteins 3) After translation, different proteins are formed which are needed for the assembly With Dengue vs. Zika virus of viral particle Need to delineate which is which since the same vector EX. M protein, E protein, NS1, Proteases (NS2b), NS4a (1) Dengue Virus AUG start codon | 5’ to 3’ untranslated region | stop codon → viral genome (+SS RNA) → Enveloped, with viral glycoproteins having hemagglutinin activity translation to protein → long polypeptide (amino acid chain) - - host + viral NS2B/3 protease - - > ○ Hemagglutinin in a virus - a clumping factor that clumps the RBCs or erythrocytes structural & nonstructural proteins → assembly of viral particle Icosahedron capsid Genome Structural genes (proteins needed for Non-structural genes SS RNA (+) strand assembly of viral particles): Non-structural proteins: ○ Mechanism of (+) RNA strand: Genetic material is mRNA itself that the virus can use (1) Core protein (capsid) NS1 for translation (Recall: Class IV Baltimore Classification) (2) *M protein NS2a (3) *P or E envelope Protein NS2b (protease) NS3 (protease) *2 & 3 Form the outer structure of NS4a virion NS4b *NS5 – needs RNA-dependent RNA Genomic RNA (at the center) → CMEE: Capsid protein → M protein → E protein → Envelope polymerase (RD RNA proteins poly.) ○ If you have (+) SS RNA, RD RNA poly. can synthesize the (-) stand from it These are ALL proteases/enzymes that can (Above image) Viral genome: With upstream → downstream of genome degrade proteins 1) 5’ Untranslated region (AUG = Start codon) → 3’ Untranslated region (Stop codon = stopping the protein translation) Serotypes 60-75% identity at amino acid level A new serotype, described in Somewhere in between the untranslated region, we have the whole frame of DEN-1 Malaysia the gene that will encode the structural & nonstructural proteins The difference in the DEN-2 DENV5 EXTRA NOTE: serotype is based on the DEN-3 ○ In the dengue virus, it's a long base where genes that are close to protein sequence, DEN-4 Because of different MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY amino acid level serotypes, the dengue vaccine can be tetravalent against the 4 serotypes Protein E: Main Viral 1. Hemagglutinin - for binding virus to host cell Protein Hemagglutinin - elicits an immune response in the host as it is the antigenic determinant (recognized region = binding) 2. Fusion with endosomal membrane at ↓lower pH (acidic) 3. The main antigen component of the virus RNA genome ○ Where does the virus usually replicate in the cell? → Cytoplasm; within intracellular vesicles Receptor Binding w/ hemagglutination (protein E)→ Virion-Endosomal Membrane Fusion @↓pH → @Cytoplasm Uncoating = +SS RNA → Translation → Replication & Assembly in Intracellular Vesicle → @Golgi App. Glycoprotein Maturation & Synthesis → Fuse with Plasma Membrane → Release 1) Dengue virus, by means of Protein E (= hemagglutinin), binds to the host cell through receptor binding For the longest time, there were only four serotypes. After 50 years, a new dengue Virus → binds to specific receptors present in the host plasma membrane serotype was reported in Malaysia 1) Enters the cell through fusion: forming the viorion-endosome membrane fusion During that time, it caused an outbreak. When they checked, the cause was not one of the Drop in ↓ pH ( = acidic) + receptor-binding → promote the formation of four serotypes, but rather a new one. endosome/virion conformational change → membrane fusion - - - - - - - - 1&2 - binding & fusion = endocytosis - - - - - - - - 2) Uncoating → release of viral genome (+SS RNA) in the cytoplasm 3) Translation occurs in the cytoplasm (upon release of the genetic material from the endosome) Unlike DNA, no need for transport into ✖ nucleus pa for DNA replication → ✖ transcription. For dengue, deretso na translation as the genetic material (+) SS mRNA is the template for translation already In the cytoplasm, (5) the flaviviruses (degnue) replicate & assemble (partially) within the intracellular vesicles → (6) golgi apparatus for maturation & synthesis of glycoproteins → (7) fuse with plasma membrane and virion release 8) Membrane associated RNA replication MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY It acquires its envelope in the plasma membrane Hemorrhagic Fever” Transmission May lead into “Dengue Shock Syndrome” 1. Intravascular plasma leakage into interstitial spaces due to inflammatory vasoactive cytokines 2. May experience hypotension (low blood pressure) 3. Circulatory shock may happen 4. Organ system impairment may occur (CNS impairment, acute There are a lot of cases of dengue in the PH kidney damage, liver injury) General Transmission of Dengue: Wild healthy aedes aegypti (does NOT harbor virus yet) → (1) bites an infected person → (2) blood is ingested by the mosquito = dengue infected → incubation time takes 8-10 days for dengue virus → (3) dengue infected mosquito bites another healthy person → (4) that person gets dengue 4-13 days later Febrile Phase Critical Phase (3-7 days) Recovery Phase 1. Retroorbital pain (pain Occurs after the fever 1. Decrease in ↓vascular behind the eyes) has subsided – lasts for permeability = no more 2. Nausea & Vomiting 24-48 hours leakage 3. Rashes 1. Low↓ platelet count 2. Vital signs beginning to 4. Occasional abdominal (thrombocytopenia) normalize pain 2. Vomiting of blood (hematemesis) No fever + normal BUT still 3. Blood in the stool have weakness (melena) 4. Nose bleeding (Epistaxis) 5. Notice puro General Idea: When you get infected with dengue viruses, you have a lifelong immunity (in blood-related^ = “Dengua any of those 4 serotypes). But there was an event (2005-2012) in Nicaragua, that DENV1, MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY DENV2, DENV3 can have homotypic infections = where you can get dengue again if you are exposed to a specific serotype AGAIN ○ Nag ka dengue ka na before then exposed nanaman with the SAME serotype = your body memory B cells promote the synthesis of new antibodies specific for the serotype Homotypic - exposed to 1 serotype, you have antibodies for this type = long/lifetime immunity BUT you can get reinfeced mas mild/asymptomatic ○ With dengue infections, it is an immunological type of infection; the antibodies synthesized in the body are highly specific when you are infected in any of the serotypes TLDR: You can have homotypic infection = lifelong immunity = exposed to same OR different serotype previously = dengue virus reinfection ND1 ka nagka-dengue tapos exposed ka to another serotype = magkaka-dengue ka pa rin betch Dengue hemorrhagic fever or shock syndrome Pathogenesis Upon human inoculation (pumasok yung virus sa katawan) → virus initially May occur in individuals (usually children) with pre-existing non-neutralizing heterologous replicates @skin cells, such as keratinocytes and Langerhans cells (local dengue antibody against dengue with a different serotype dendritic cells: phagocytes and antigen-presenting cells = MHClass) → ○ Passively acquired maternal antibodies enters macrophages → entry into the bloodstream → myeloid lineage ○ From a previous infection (Lymphatic System ) Kapag nasa bloodstream na, viremia starts = starts fever na Situational Example Dengue viruses primarily infect cells of the myeloid lineage, Yung nanay infected with dengue virus serotype 1 (DENV1) = mom has antibodies directed including macrophages, monocytes, and dendritic cells. There is for its serotype. Mother can pass the specific antibodies for only DENV1 to the baby. evidence of infection of hepatocytes (liver) and endothelial cells. ○ If nagkaroon ng dengue yung baby is caused by another serotype, DENV3, the effect Haematogenous spread is the likely mechanism for seeding of on the baby is different. peripheral organs and the occasionally reported infection of the If you have antibodies from previous, these are non-neutralizing antibodies specific to 1 central nervous system. serotype meaning they canNOT neutralize other serotypes and worse could even aid in its Spread - virus is disseminated to other parts of the body entry to immune cells + increase viral replication. Leading to a more profound response = Dengue shock syndrome → (continue image below) MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY TLDR: Body can have a different effect if infected with another serotype that does not match neutralization test sensitive as molecular the pre-existing antibody, as the antibody is directed for the previous serotype only. (PRNT) tests during the first 0-7 days of symptoms. Diagnosis of Dengue After day 7, NS1 tests are NOT recommended Laboratory Test Description 1. Dengue NS1 Requested between 1-5 days of illness (Non-structural Used to detect acute (sudden onset) phase of dengue Protein) Combination RT-PCR and NS1 tests: offer earlier and more specific diagnosis (80-90% 2. Dengue IgM/IgG Requested beyond >5 days of illness sensitivity if assessed 1-3 days after the onset of illness) and are considered virological proof of IgM 1st → IgG Used to detect the presence of IgM and to determine previous infection (direct virus detection) later dengue infection with the presence of IgG Disadvantage: may cross-react with Zika infection or other Treatment Supportive flavivirus infection Paracetamol ○ If positive, it does NOT necessarily mean that youre Increase fluid intake infected with dengue. You have to rule out that its not ○ As there is a vascular permeability that can lead to zika or other flavivirus infection leakage thus dehydration 3. ⭐Polymerase The gold standard to confirm dengue infection Bed rest Eat nutritious foods chain reaction (PCR) ✖ Ibuprofen or aspirin should NOT be taken due to risk of bleeding ← anticoagulation properties 4. Other tests: Routinely used in the hospital as standard dengue diagnostic Tinatake kasi siya kapag may fever which is dapat HINDI Total WBC Count test Platelet Count Decreasing WBC, decreasing platelet count and increasing Prevention Use mosquito repellant Hematocrit hematocrit (RBC) are indicative of dengue infection Wear protective clothing ↓WBC + ↓platelet +↑hematocrit (RBC) = dengue infection Regularly remove ✖ stagnant water at home ○ Vector can breed in stagnant water Laboratory Diagnosis of Dengue Use a bed net Secure windows and doors at home Serologic Tests: Molecular Method: Immunologic Method: MAC-ELISA & PRNT RT-PCR NS1 Test ( genes encoding the prM and E proteins of the corresponding genes of the 4 wild-type dengue viruses – replaced with genes corresponding to the 4 dengue serotypes = tetravalent (Evidence based) As suggested by WHO, dengvaxia should ONLY be given to those who had dengue previously. This is because of an increase of incidence hospitalization and severe dengue illness in children that NEVER HAD dengue. ○ As per WHO, possibility for negative (0%) people (those with NO antibodies against the serotypes), vaccination may be ineffective or may theoretically increase future risk of hospitalization or severe dengue illness. ○ Advice: Give the doses to those who were infected previously. Kwento ni miss: Maraming nanay na vaccines are not safe for kasi nag ka issue yung dengvaxia, nakakamatay daw. Incidence of death may happen but VERY low. Dahil sa issue na to, after years, nag ka polio sa pinas kasi maraming di nakapag vaccine. MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY (2) Japanese encephalitis virus - replication cycle similar to dengue virus Characteristics SS (+) strand RNA Enveloped, icosahedron capsid Mainly found in some parts of Asia Affect children >> more than adults Vector - Culex tritaeniorhyncus How can one explain the excess cases of severe dengue in the vaccinated seronegative population? ○ Hypothesis: If you are sero-negative, the vaccine can initiate an immune response to dengue that predisposes them to a HIGHER severe risk. Vaccine acts like a primary-like infection and a subsequent infection with the 1st wild type dengue virus is then a “secondary-like” clinically more severe Main cause of encephalitis in Asia infection ○ Involve infections of neurons in CNS = lead to neurologic The vaccine does NOT CAUSE excess cases but instead induces immune status with INCREASED infections RISK to greater severity of subsequent infections Childhood neurological infection TLDR: Immune response, initially, is due to the vaccine. Subsequent infection, if nakagat ka = May cause permanent brain damage secondary infection will be a SEVERE type Mosquitoes infected by biting infected pigs Humans are the dead-end hosts = NO longer ✖ transmit to other hosts kapag nasa human na Refer the the figure above: ○ If unvaccinated → exposed = primary infection → exposed to another serotype = secondary infection → post secondary infection ○ If vaccinated with NO previous infection → body advances ✖ primary na, SEVERE infection (predisposed yung katawan) → secondary like = RISK is high ○ If vaccinated with previous infection → antibodies are ready from the primary infection → vaccinate → gets exposed again to dengue = NOT SEVERE; ✖ high risk– remember kaya ito lang recommended for dengvaxia Generalized transmission cycle of mosquito-borne (flaviviruses) causing enchepalititis^ MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY Amplification cycle Diagnosis ELISA-serum / CSF JEV-specific IgM antibody – serologic ❖ Culex mosquitoes → can infect birds (✔common host) AND pigs (✖ not common ○ In IgM antibody, you have to rule out dengue as it can host) cross-react in tests Culex mosquitoes bites an infected (1) pig → mosquito is infected → bite the MRI for brain lesions (2) bird → bird is infected → if it bites a (3) human, person is infected = suffer from disease BUT it does NOT contribute to the transmission or Treatment None ~ba yan.. multiplication of the virus Supportive Paracetamol In japanese encephalitis virus, it has overwintering mechanism to survive long periods of time or Increase fluid intake extreme conditions → like being dormant Bed rest Eat nutritious foods 1) In tropical environments, it can establish overwintering maintenance = can survive BUT NOT multiply ~weird tropical pero over(wintering) Prevention Use mosquito repellant 2) In hibernated vector = EX. animals hibernating over winter, virus can be supported through Wear protective clothing this Regularly remove stagnant water at home 3) Prolonged viremia - in a certain host for long periods of time Use a bed net 4) Alternative vector Secure windows and doors at home 5) Vertical transmission **Notice same lang Supportive & Prevention kay dengue Pathogenesis Infected mosquito bite → blood or macrophages → enters the Vaccine Japanese Encephalitis (JE) vaccine given in a two-dose series; blood brain barrier → brain (may also affect the basal ganglia, given 28 days apart ~ VS dengue 3 doses every 6 months thalamus, midbrain, & pons) ○ Inactivated vaccine = virus is inactivated to weaken the virulence Clinical features Asymptomatic to life threatening Given in adults and children Incubation period 4-14 days (WHO) A booster dose should be given (3rd dose) if a person has 1. High grade fever received a 2-dose primary vaccination or if there is a risk for 2. Headache re-exposure 3. Neck stiffness 4. Nausea & vomiting (3) Zika Virus 5. Disorientation – Especially if the virus is in the brain Guillain-Barre Syndrome (autoimmune) + Transplacental + Microcephaly 6. Coma 7. Seizures Enveloped 8. Spastic paralysis Icosahedron capsid 9. Death SS RNA (+) strand **Pare-parehas lang sila flavivirus Complications Survivors suffer from: Neurologic impairment Transmission Bite of infected Aedes aegypti, Aedes albopictus ~dengue twin Behavioral problems Sexual intercourse Speech deficits Correlates with higher incidence of Guillain-Barre syndrome: Paralysis ○ Autoimmune disease (self attack self) – Own immune Recurrent seizures system attacking own nervous system NS → nerves are MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY destroyed → can lead to paralysis of whole body Family Retroviridae Transplacental transfer for at least 3 months after infection ○ Mother to baby; birth Genus Lentivirus Infection of fetus (still developing) - microcephaly and other (1) Human Immunodeficiency Virus (HIV) congenital abnormalities ○ Brain NOT fully developed Common human virus ○ Possibly due to vertical transmission: infected mother 2 strains: transmits virus to the baby/fetus ○ HIV 1 - 99% of global cases ○ Microcephaly: death of progenitor brain/neural cells ○ HIV2 needed for full development of the brain OR NS Possible HIV+ = infected by virus but NOT yet aids A “syndrome” → Weak immune system thus susceptible to diseases due to being HIV+ Enveloped ○ Envelope with proteins like transmembrane proteins + nucleocapsid + genetic material (2 copies of +SS RNA 5’ → 3’) ○ 3 Genes - GAGS PRIE ENV Gag genes (2 vertical lines) = code for structural proteins of HIV Pol genes = code for reverse transcriptase & integrase Env gene = code for envelop proteins Helical nucleocapsid within an icosahedron capsid 2 identical copies of (+) SS RNA 5’ → 3’ ○ Despite starting with +SS RNA this is NOT used directly as mRNA for protein translation = class VI retroviruses Class VI retrovirus ○ Transcription of (-) DS DNA in reverse transcription by reverse transcriptase (+) SS RNA → cDNA (complementary); 3’ to 5’ so the referred (-) DS DNA^ → DS DNA (intermediate) 5’ to 3’ → 1 strand used → transcription to mRNA MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY Viral Enzymes in virion @nucleocapsid 3: reverse transcriptase, integrase, protease 1. Integrase – integration of viral material into host 2. Protease – enzymes degrading other proteins/enzymes 3. Reverse transcriptase (has OWN) – mechanism of mRNA transcription, conversion of RNA → cDNA (complementary) Host remain chronically infected Within the macrophage, HIV replicates → makes an altered form of gp120 that recognizes a different coreceptor, CXCR4, on Th cells → HIV virions are released from macrophages and proceed to infect and replicate in Th lymphocytes; Th cells that produce HIV NO longer divide and are eventually diminished by attrition Class VI SS RNA (+) = HIV ○ From cDNA another strand is synthesized to form a dsDNA intermediate → Transcription of (-) strand = mRNA Target cells: Cells with CD4 receptor – common denominator sa main receptors (pair) ○ HIV can really replicate in antigen presenting cells = macrophages/phagocytes + lymphocytes (T-helper) + other immune cells HIV transmembrane with specific receptor proteins; proteins allowing binding to receptors in the host First infected: macrophages receptors - CD4 and CCR5 ○ Followed by: T helper (Th) lymphocytes receptors- CD4 and CXCR4 MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY Mechanism: 7. New viral RNA synthesized = new/starter genomic material to make viral proteins 1. HIV transmembrane structure spike proteins with gp120/gp41 (proteins on envelope 8. Assembly & packaging of viral particles of virus) 9. Exit cells via lysis 2. Gp120/gp41 (HIV) binds to receptor cells of macrophages CD4 + coreceptor CCR5 HIV transmembrane spike proteins gp120/42 bind to receptor cells @macrophages CD4 + CCR5 ○ Receptor-coreceptor pair as the main receptors: CD4 + CCR5 = Fusion of → Fusion → Nucleocapsid Entry → Nucleocapsid Uncoating → RT converts mRNA to DNA → viral HIV envelope to host cell (membrane) DNA integrates host genome DNA → New viral RNA → Assembly & packaging → Lysis exit ○ Within macrophage: Replication HIV can replicate = new altered gp120 protein conformation that recognizes/binds on co-receptor CXCR4 on T lymphocytes – only occurs IF replication happened Macrophages → it reaches T helper lymphocytes → proceed replication from mRNA → cDNA → DNA (integrated in host DNA) Proteins are specific so it cannot just bind Once in Th lymphocytes, HIV does NOT divide anymore = since masisira rin T lymphocytes eventually/go back to og mechanism targeting macrophages ○ HIV spike proteins w/ gp120/41 → bind to macrophage w/ CD4 + CCR5 main receptor-coreceptor pair → Fusion virus envelope + host cell membrane → New gp120 bind to Th lymphocyte w/ CD4 + CXCR4 → Stop division → back to macrophages (+) SS RNA - - Reverse Transcriptase - - > cDNA (complementary) → DS DNA (intermediate) → 1 ○ Infection to others: DNA strand used → transcription to mRNA After replication in T lymphocytes → the new viral particle synthesized & spread would only have the og co-receptor CD4 + CCR5 NOT the modified for CXCR4 – depende pa rin sa genome type ng virus 3. Fusion → allow entry of nucleocapsid (genetic material + reverse transcriptase + integrase + other proteases) to cytoplasm of the cell 4. Nucleocapsid enters cell → uncoating 5. RT converts mRNA of genetic material 5’ → 3’ to viral DNA ○ mRNA → cDNA → viral DNA (intermediate) 6. Viral DNA integrate host genome (DNA) via inegrase ○ What makes vaccine design against HIV tricky is that virus DNA can be integrated into host DNA = immune system canNOT read/identify/attack In chromosome; dormant stage = provirus the virus → NO approved vaccine, only anti-retroviral (Prophylaxis) that ○ Person infected shows NO signs of symptoms – HIV+ ka na pala di mo alam prevent replication of virus – taken with potential exposure = preventing viral Virus in host DNA → begins to replicate & kill other cells = surprise replication immunocompromised Taken within 48-72 hours (2-3 days) to halt viral replication for HIV progression = T lymphocyte decline some time = virus eventually degrades in system ○ EX. CD4 count of 2-4 nalang gg MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY ○ Can be latent for long periods ○ Avian flu A virus H1N1 (-) SS RNA, 7-8 different segments Enveloped ○ Contain antigenic properties: surface projections/proteins – hemagglutinin or neuraminidase activity → determine antigenic variation of influenza viruses and host immunity Helical symmetry Particles: round or filamentous Receptor on host-cell-sialic acid (NANA) in plasma membrane molecules in glycoproteins & glycolipids of host membrane Hemagglutinin – binding Neuraminidase – mucin layer → epithelial cells Binds virus particles to susceptible cells Sialidase → bind to receptor sialic acid Highly immunogenic, major antigen from host membrane against which neutralizing antibodies are Removed terminal sialic acid from directed cellular, the viral surface glycoproteins 1) Release virus particles from the cell 2) Prevents aggregation of virions HIV+ individuals become susceptible (kasi nga Th lympho. declines) to many infections like Helps the virus negotiate through the fungal (EX. yeast) kaya immunocompromised talaga sila mucin layer in the respiratory tract to ○ Low CD4 count = weak immune system – EX. candidiasis → ear infection naman next reach the target epithelial cells Laboratory Diagnosis Detection of anti-HIV Detection of HIV load antibody response can require a period 1. RT-qPCR: monitor progression of an HIV of 6 weeks or more following infection infection and the effectiveness of 1. Screening: ELISA or EIA chemotherapy 2. Confirmatory: Immunoblot ○ Quantify expression of genes 3. Confirmatory: Indirect immunofluorescence Family Orthomyxoviridae - Influenza Virus See typical glycoplipid with NO charge VS host membrane with (-) charge due to sialic Genus Orthomyxovirus acid receptor component ○ NANA (-) sialic acid ng host fascilitates binding of the virus Flu viruses: ○ Spanish flu: 1918 pandemic MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY Antigenic drift - mutation overtime accumulates = surface antigen change ○ Virus highly mutating; prone to mutation as they are dependent on host mechanism, accumulation of mutations overtime = changes in surface antigens Antigengic shift & drift = every year may new strain of influenza → need yearly flu shots ○ Vaccines against influenza today may NOT be effective in 2 years time kasi baka bago na naman properties niya 1) Influenza type A (human and animal hosts): highly antigenically variable, responsible for Genome most cases of epidemic influenza (-) sense, SS RNA → virion with RNA-dependent RNA polymerase OR “RNA replicase” 2) Influenza type B (human host): may exhibit antigenic changes, sometimes causes epidemics ○ (-) sense is converted to (+) sense 3) Influenza type C (human and swine hosts): antigenically stable and causes only mild illness Linear, segmented (8 segments) in immunocompetent individuals ○ segmented nature of the viral genome permits genetic reassortment when 2 influenza viruses infect the same 1 cell → mixtures of parental gene segments may be assembled into progeny virions → may result in sudden changes in viral surface Family Picornaviridae - Poliomyelitis Virus antigens → can result in antigenic drift, antigenic shift → high rate of natural Naked, icosahedron variation among influenza viruses SS (+) sense RNA Viral reassortment and transmission from other species is thought to explain the 12 genera emergence of new human pandemic strains of influenza A viruses Genetic reassortment: 2 influenza infect 1 cell = parental gene segments + progeny virion mix assembled → sudden viral surface antigen change = antigenic drift, shift → variation Antigenic shift – shared genomic property (RNA in host) = influenza new type + surface ○ 2 infects 1 – shared genomic properties (genetic reassortment); combining RNA in the host = new progeny with shared genomic properties (new type of influenza virus) with new (antifenic surface) MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY Transmission: fecal-oral route ○ EX. buffets no longer self-service = importance of food handling etc. MORE TYPES 1. Hepatovirus 2. Kobuvirus 3. Parechovirus Hepatitis A virus Aichi virus parechoviruses 4. Cardiovirus 5. Aphthovirus Cardioviruses Foot-and-mouth disease viruses 2 Vaccines 1) Formalin-inactivated vaccine (Salk) – dead for immunodeficient/compomised ✖ antibodies ○ Inactivated = killed virus itself (dead) ○ Will NOT allow body to initiate antibodies in gut (duodenum) NO nasal & duodenal IgA antibodies – lower secretory antibodies 2) Oral live attenuated vaccine (Sabin) – weakened used to eradicate polio ✔ antibodies ○ Weakened viruses, still alive = virulence is also weakened High; all antibodies present ○ ⭐ Winner with life long immunity Though studies show that the virus still persists in individuals in small Naked: once enters cell → RNA used for protein translation for proteins (used in viral numbers particle assembly) ○ If you are ✖ immunodefficient/compromised – this is NOT suggested All of the replication occurs in the cytoplasm = di na need sa ✖ nucleus – Class IV Body may respond differently = overproduction of inflammatory cells & cytokines Enterovirus — (1) Enteroviruses; (2) Rhino viruses Antibody is the major protective immune response to enteroviruses ○ Secretory antibody: prevents initial establishment of infection in the oropharynx Enteroviruses – resistant Rhino viruses – sesitive and GI tract 1. Polioviruses 1, 2, 3 Naked, icosahedral ○ Serum antibody: prevents viremic spread to the target tissue and therefore 2. Coxsackie viruses a and b At least 100 serotypes disease 3. Echoviruses Major causes of the common cold Cell mediated immunity is NOT usually involved in ✖ protection but may play a role in Capsids are very resistant to harsh Sensitive to acidic pH resolution and pathogenesis conditions in the environment (sewage Replicate poorly at room temperature systems) and GI tract (low pH) above 33°C → usually limit rhinoviruses Entry → GI tract → blood stream = 1st set of to causing only upper respiratory tract viremia → other parts of body infections MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY Family Rhabdoviridae - Rabies Virus Genus Lyssavirus *SKL: not just animals, pati plants Unique bullet-shaped, enveloped with glycoprotein spikes SS (-) RNA → virion with RNA-dependent RNA polymerase – Class V You do NOT stop sa (+) strand RNA – need pa rin synthesize ang (-) strand RNA as this is the genetic material that must be replicated MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY Enveloped, with longest SS (+) RNA Helical nucleocapsid Glycoproteins on envelope: allow the virus to endure the conditions in the GI tract, can be spread by the fecal-oral route Most infected: bats > cats > dogs ○ EXAMPLE Virus can replicate in our muscles → PNS → brain → NS affecting brain stem, cerebellum, Severe acute respiratory syndrome coronavirus (SARS-CoV) in 2002 other parts of the brain = impared brain function; hallucination Middle East respiratory syndrome coronavirus (MERS-CoV) in 2012 ○ If you get bitten → inject a lot kasi rabies disease progression takes time ~10 SARSCOV 2 in Dec 2019 years before worse talaga Human coronaviruses - 33°C to 35°C optimum temperature for viral replication, cause You can be infected but long disease progression – need pa booster shots URTI Aggregates of viral nucleocapsid = Negri body Animal coronaviruses - 37°C, cause systemic disease in humans SARS-COV 2 Coronaviridae Genus Betacoronavirus Coronavirus Causes coronavirus disease 2019 (COVID-19) Shares 79% genome sequence identity with SARS-CoV and 50% with MERS-CoV Coronavirus from bat Rhinolophus affinis from Yunnan province China, full-length genome sequence 96.2% identical to that of SARS-CoV-2 Coronavirus from Malayan pangolin smuggled from Southeast Asia into Southern China from 2017 to 2019: 92.4% sequence similarity Envelop with spike protein specific for ACE2 ○ SARS-CoV-2 receptor in human host: angiotensin-converting enzyme 2 (ACE2) (same as for SARS-CoV) MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY SARS-CoV-2 also recognizes ACE2 from pig, ferret, rhesus monkey, civet, cat, pangolin, rabbit and dog Viral Spike (S) protein: requires cleavage with participation of host proteases to activate the entry of the virus via the endocytic route GeneXpert ○ For SARS-coV-2: quantifying gene expression - like qRT PCR Lateral flow immunoassay For COVID-19 using SarsCoV2 EX. COVID-19 Rapid POC CE-IVD kit – Serological, we take blood sample Detects IgG and IgM antibodies specific for SARS-CoV2 from whole blood, serum or plasma ○ If you are infected → produce antibodies IgG, IgM – more abundant in blood Lateral movement: left → right Therapeutic agents inhibits: Receptor-binding, Fusion, Replication Baltimore classification: Class IV I. Sample Pad (load sample) → both against SarsCoV2 Mechanism 1) Patients IgM 1. SARS-coV-2 spike protein → bind to receptor ACE2 2) Patients IgG 2. Facilitate fusion & entry II. Cojugation Pad - have Ig antigens directed for the antibodies 3. Nucleocapsid in cytoplasm → release genetic materia AuNP-COVID 19 antigen +IgM 4. Read RNA to translation AuNP-COVID 19 antigen + IgG 5. Synthesis of proteins → form final viral particles AuNP-rabbit IgG (control) Spike protein → Bind receptor ACE2 → Fusion → Nucleocapsid Entry → Cytoplasm → RNA Blood sample (w/ IgG + IgM) → 1 sample pad w/ your antibodies → 2 conjugation pad w/ antigens → translation to proteins → Final viral particles MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY antibodies (IgG & M) bind to antigen (2 for G & M) III. Nitrocellulose Membrane: 3 regions – binds to complexes formed from conjugation pad ***All regions have color*** 1) M region: anti-human IgM BINDS (AuNP-COVID-19 antigen +IgM) 2) G region: anti-human IgG BINDS (AuNP-COVID-19 antigen + IgG) 3) C region: anti-rabbit IgG BINDS (AuNP-Rabbit IgG); rabbit control A coloured line should ALWAYS appear in the control line region, indicating that the proper volume of specimen has been added and membrane wicking has occured – if kulang volume, NOT enough antibodes to bind = false (-) negative Lipid nanoparticle mRNA → intramuscular injection → cytoplasm → translation to spike proteins → immune response → T helper cells → cytokines & interleukin + memory cells + kill infected cells mRNA in a form of lipid nanoparticle injected intramuscularly → mrna goes to the cytoplasm → ribosomes read & translate mRNA to proteins ○ mRNA = sequence of spike proteins – body synthesizes spike protein eliciting immune response against this T helper cells produce cytokines, interleukins that will also stimulate more T cells to make more memory cells + kill infected cells 3 SARSCOV2 Vaccines 2) Viral Vector Vaccine Engineered weak chimpanzee virus w/ DNA coding spike proteins + DNA vector template 1) mRNA Vaccine Lipid nanoparticle mRNA + T helper cells MDMICRO MEDICAL MICROBIOLOGY (LECTURE) TERM 3 AY. 2023-2024 4TH LONG EXAMINATION | DR. LLEWELYN ESPIRITU | RNA VIRUSES TRANSCRIBED BY: ANGELINA ARQUINES & JAZMINE TANHUY 3) Subunit Vaccine Removal & extract spike protein + insertion for multiplication Also used in adenovirus Remove & get (gene cloning/expression) spike protein → insert in bacteria, yeast, animal cells that multiply → more spike protein copies → spike proteins extracted → formulated as vaccine injected → spike proteins recognized by immune system = elicit human response DNA encoding spike proteins → engineered, modified into weakened chimpanzee virus = commercially available vaccine → injected in body → DNA vector template in human cells → DNA released for replication, transcription, & translation → synthesized (by ribosomes) spike protein → recognized & elicits immune response = antibodies against COVID-19