8.SARS COV-2.pptx LAB DIAGNOSIS PROPHYLAXIS

FOR MEDICAL

1. SARS COV- 2 Pathogenesis, Laboratory Diagnosis, Management and Prophylaxis

2. OUTLINE Introduction Pathogenesis Laboratory diagnosis Management Prophylaxis

3. Introduction SARS-CoV-2,the causative agent of COVID-19, a newly emergent coronavirus. F irst detected in Wuhan, China, in December 2019. O rder Nidovirales , Family Coronaviridae , Subfamily Coronavirinae , genus coronavirus 4 genera( α coronavirus, β coronavirus, γ coronavirus, δ coronavirus) SARS CoV 2 strain comes under the genera β coronavirus subgenus sarbecovirus .

4. In December 2019, China reported an outbreak of pneumonia of unknown causes in Wuhan, the capital city of Hubei province Earlier, 2019–novel Coronavirus (2019–nCoV) O n February 11, 2020, ICTV renamed as SARS–CoV2 ( genome resembled to SARS- CoV ) WHO named the resultant disease as Coronavirus disease (COVID-19 ) WHO designated SARS-CoV-2 a Public Health Emergency of International Concern on January 30, 2020, and pandemic on March 11, 2020 Epidemiology

5. On January 11, the first case was reported outside mainland China in Thailand . W ithin months, the disease spread to all the continents except Antarctica.  India reported its first case on January 30, 2020 in Kerala. The first COVID-19 related death in India was reported on March 12, 2020.  By the second week of April, the disease spread to all states in India except Sikkim. Pandemic occurred in 3 waves, first around September 2020,second in April 2021, third in January 2022.

6. On 25 th March 2020, Govt of india declared a country wide lockdown till 31 st march, which was extended upto 14 th April 2020, Further extended to 3 rd may then upto 17 th may 2020- covers 4 phases of lockdown f/w gradual un-lockdown in the country, spread in 6 phases(one month each) upto 30 th november2020. Aarogya setu app: Launched by the GOI on 2 nd April 2020(location based surveillance app)

7. Morphology Enveloped, spherical , 120-160-nm, G enome - unsegmented +ss RNA(27–32 kb), surrounded by helical nucleocapsid protien 9–11 nm. 20nm long club or petal shaped projections, widely spaced, giving appearance of solar corona. 4 structural proteins: S,E,M and N 16 Non-structural Proteins, - P roteases (nsp3 and nsp5) and - RdRp

8. Variants of SARS-CoV-2 : (Variants of concern) B.1.1.7 (Alpha)- F i rst detected in the U.K. in September 2020, N501Y mutation in the RBD region of the Spike protein. E nhance binding b/ w the SARS-CoV-2 Spike protein and the human ACE2 receptor,  2) B.1.351 (Beta)- Arose in South Africa, both the E484K and N501Y mutations. 3) P.1 (Gamma)- E merged in Brazil; detected in the USA by the end of Jan 2021. T hree (K417T, E484K, and N501Y) within the RBD of Spike. E484K- affect the ability of antibodies to recognize and neutralize the virus.

9. 4) B.1.617.2 (Delta): “double mutant” variant, First identified in India in December 2020 (second wave) L452R- S tabilize the interaction between the spike protein and the host cell -increase infectivity E484 mutation, higher infectivity and transmission rates. 5) B.1.1.529 (Omicron) : Have high transmissibility but produced less severe symptoms and hospitalization. In India 3 rd wave was largely attributed by this variant.

10. Mode of transmission- By droplet borne infection, either directly or indirectly, through fomites. Airborne transmission can occur during medical procedure. R isk factor- - Advanced age -Male sex - Preexisting comorbidities - Elevated body mass index - Substance use, such as alcohol, opioid, or cocaine use disorder, and current or former smoking both increase risk. - Pregnant women Pathogenesis

11. Host Cell Entry

12. C ourse of infection goes through the following stages – - Viral invasion and replication, - D ysregulated immune response, - M ultiple organ damage and recovery.  V irus enters the host cells, replicates, assembles and is released extracellularly to target cells, damage and destruction of alveolar epithelial cells. At the same time,large number of PAMP and DAMP molecules are released - stimulate the innate immune response, release large quantities of cytokines, chemokines, proteases and free radicals, ARDS, sepsis and MODS

13. Clinical presentation- IP- 2 to 14 days . Fever is probably one of the most common D ry cough A nosmia or hyposmia and loss of taste  Anorexia Myalgia and dyspnea Gastrointestinal symptoms included diarrhea, abdominal pain, and vomiting/nausea 

14. Clinical Severity: Asymptomatic Mild disease/ILI ((influenza-like illness) - H ave fever, cough, sore throat , malaise, headache. Moderate disease/Pneumonia- ILI plus any one: 1) Respiratory rate ≥ 24/min, breathlessness 2) SpO 2 : 90% to ≤ 93% on room air S evere disease/SARI/Severe Pneumonia-Pneumonia plus any one: 1)Respiratory rate ≥ 24/ min,breathlessness 2)SpO2: 90% to ≤ 93% on room air Critical- R espiratory failure, Septic shock, and/or Multiorgan failure

15. Laboratory diagnosis Types of specimen- Nasopharyngeal specimen is a preferred choice -O ther specimen- O ropharyngeal (OP) specimen,Nasal mid-turbinate (NMT)swab, A nterior nares (nasal swab; NS) specimen and N asopharyngeal wash or N asal aspirate (NA) specimen , T hroat swab, BAL, E ndotracheal as p irate can be collected alternatively.   For serological assay- Blood sample

16. Placed in a tube containing viral transport media(VTM) I f the shipping of specimen to the reference diagnostic laboratories required, the sample must be transported in triple packaging system; Sample vial must be properly labeled and sealed and kept in outer covering of absorbent material(Primary container) Secondary container should be placed with frozen gel packs in thermocol box.

17. NAAT: Real time RT-PCR- Gold standard Require biosafety level-2 facility Gene targets for screening – Spike protein (S), Envelope protein (E), Membrane protein (M), Nucleocapsid protein (N) Gene targets for confirmation - RNA-dependent RNA polymerase ( RdRp ), Open reading frames (ORF1a/b), N2 nucleocapsid Automated real-time RT-PCR (CBNAAT and Truenat )

18. 2)Antigen detection assay: Point-of-care test; detects nucleocapsid protein antigen in nasopharyngeal swab 3)Antibody (IgG) detection assay: Used for serosurveillance and survey in high-risk and vulnerable group; not for clinical diagnosis 4)Sequencing: To determine mutations in the viral genome 5)Viral culture: Used for research purpose - Require biosafety level-3 facility 6)Nonspecific tests include: Radiology (chest CT scan): Ground-glass appearance Biomarkers: IL-6, D-dimer, Elevated serum ferritin, Elevated C-reactive protein

19. Course of the diagnostic markers in COVID-19 .

20. 7) LAMP TEST(Loop Mediated isothermal amplification): Developed by Tsugunori Notomi at Eikens chemicals, Tokyo, Japan. Having higher sensitivity and specificity than traditional PCR. Approved for Covid diagnosis in India as Lume-NCoV Kits from Agappe Diagnostic Kochi,Kerala . 8) CRISPR ( Clustered Regularly Inter Spaced Short Palindromic Repeats): Developed in the USA as the ‘Sherlock’ test. The test developed in India based on this named ‘FELUDA’. Commercially launched in India by Tata medical diagnostics as ‘ TataMD CHECK CRISPR test’ Test uses CAS9 protein to find and bind to target RNA. Coupled with paper strip chemistry to allow a visual reading.

21. Management N o definitive therapy Symptomatic management- In patients with severe respiratory distress Supplemental oxygen therapy is given immediately High-flow nasal cannula oxygenation (HFNO) Non-invasive mechanical ventilation Mechanical ventilation: In patients with moderate or severe ARDS, higher PEEP (positive end-expiratory pressure) instead of lower PEEP is suggested. Management of septic shock by—vasopressors, fluid replacement by crystalloids such as normal saline and Ringer’s lactate Convalescent plasma therapy Drugs- Remdesivir, Tocilizumab, Hydroxychloroquine (HCQ)

22. Prophylaxis WHO recommends protective measures to help prevent the spread of the disease -Wearing mask - Frequent hand washing, - A voiding touching of eyes, nose or mouth, - M aintaining a distance of at least 1 m between people, - P racticing respiratory hygiene, Chemoprophylaxis- Hydroxychloroquine

23. Types of vaccines : -Inactivated or live-attenuated - Covaxin , CoroVac and BBIBP-CorV - Protein subunit based - Novavax, Epi Vac Corona - mRNA based - Moderna, Pfizer–BioNTech (BNT162b2) -Viral vector: ( recombinant adenovirus with a SARS-COV-2 gene ) - Covishield, Oxford- Astrazeneca , Sputnik V and Johnson & Johnson COVID -19 vaccine

24. Covaxin : code name is BBV152. - P repared by Hyderabad- based Bharat Biotech International Ltd. in collaboration with the National Institute of Virology (NIV) and ICMR -effectiveness - 78-81%. - administered in two doses (4-6weeks apart) by IM route Covishield: code name AZD1222, prepared by Serum Institute of India, in collaboration with University of Oxford and AstraZeneca pharmaceuticals, UK. - Effectiveness 90% - B ased on non-replicating adenovirus vector (modified Chimpanzee adenovirus, ChAdOx1) expressing spike protein. - Administered in two doses ( 12-16 weeks apart) by IM route.

25. Sputnik V, by the code name rAd26-S and rAd5-S, was developed by Gamaleya Research Institute of Epidemiology and Microbiology, which involves an interval gap of 21 days between the two doses. - Effectiveness 92.2% CoWIN (Covid Vaccine Intelligence Network)- Indian government web portal for Covid 19 vaccination registration and also for obtaining the certificate after vaccination .

26. REFERENCES: Medical Microbiology – 28 th edition , Jawetz , Melnick & Adelberg’s Ananthanarayan and Paniker’s Textbook of Microbiology – 8 th edition Park's textbook of preventive & social medicine, 26 TH edition Apurba Sanker Sastry, Sandhya Bhat Essentials of Medical Microbiology, 3 rd Edition   Kumar KSR, Mufti SS, Sarathy V, Hazarika D, Naik R. An Update on Advances in COVID-19 Laboratory Diagnosis and Testing Guidelines in India. Front Public Health. 2021 Mar 4;9:568603. doi : 10.3389/fpubh.2021.568603. PMID: 33748054; PMCID: PMC7969786. Aleem A, Akbar Samad AB, Vaqar S. Emerging Variants of SARS-CoV-2 and Novel Therapeutics Against Coronavirus (COVID-19). 2023 May 8. In: StatPearls [Internet]. Treasure Island (FL): StatPearls Publishing; 2023 Jan–. PMID: 34033342. Mukim M, Sharma P, Patweker M, Patweker F, Kukkar R, Patel R. Covid-19 Vaccines Available in India. Comb Chem High Throughput Screen. 2022;25(14):2391-2397. doi : 10.2174/1386207325666220315115953. PMID: 35293291.