Lab diagnosis-viruses.ppt
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This document discusses laboratory diagnosis of viral infections through various methods. Sample collection from different sites depends on the suspected virus. Diagnosis involves direct microscopy to look for inclusion bodies, culturing samples in animals, eggs, or cell lines to isolate viruses. Detection methods include cytopathic effects in cell culture, antigen detection by immunofluorescence, and antibody detection through serological tests. Newer molecular techniques like PCR allow for nucleic acid detection of specific viruses.
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1. Direct fluorescent antibody staining of cells from a herpes simplex lesion can detect fluorescent infected cells under a fluorescence microscope, providing a more sensitive and specific method of diagnosis than a Tzanck prep.
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Viruses are obligate intracellular parasites which means they can only grow or reproduce inside a host cell.
The primary purpose of virus cultivation:
To isolate and identify viruses in clinical samples.
To do research on the viral structure, replication, genetics, and effects on the host cell.
To prepare viruses for vaccine production.
Isolation of the virus is always considered a gold standard for establishing the viral origin of the disease
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Viruses are obligate intracellular parasites that infect all living organisms. They possess either DNA or RNA and lack organelles like cell membranes and ribosomes. Viruses come in a variety of shapes and sizes, with the smallest being 20nm and largest 400nm. They enter host cells and hijack the cell's machinery to replicate their genome and proteins. Viruses can undergo lytic or lysogenic replication cycles. Laboratory diagnosis of viral infections involves direct visualization of viruses, detection of antigens and antibodies, and molecular methods like PCR to detect viral genes.
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Introduction
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Sample collection & Transportation
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Demonstration of virus antigen
Detection of viral growth in cell culture
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Viruses are obligate intracellular parasites which means they can only grow or reproduce inside a host cell.
The primary purpose of virus cultivation:
To isolate and identify viruses in clinical samples.
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To prepare viruses for vaccine production.
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Animal inoculation
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3. The document also summarizes the clinical presentation and pathogenesis of numerous viral infections including herpesviruses, adenoviruses, parvovirus B19, human papillomavirus, hepatitis viruses, arboviruses, and enteroviruses.
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This document discusses techniques for cultivating viruses. It explains that viruses require living host cells to replicate and describes three main cultivation methods: animal inoculation, embryonated egg inoculation, and tissue culture. Animal inoculation involves infecting animals like mice to isolate and study viruses, but it is expensive and raises welfare issues. Embryonated egg inoculation is commonly used to grow viruses by inoculating eggs at specific sites, and it is cost-effective but each virus grows at different sites. Tissue culture uses cell monolayers and is versatile but requires specialized facilities and technicians. The document provides details on each technique's advantages, disadvantages and applications.
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Lab dig virus
This document provides information on viral diagnosis methods. It discusses:
1. Direct examination methods like antigen detection, electron microscopy, and PCR to detect viruses.
2. Indirect examination using cell culture, eggs, or animals to isolate and identify viruses. Cytopathic effects and immunofluorescence are used to detect growth.
3. Serology methods like ELISA, complement fixation, and neutralization to detect antibodies produced in response to viral infection. Rising titers indicate acute infection while IgM indicates primary infection.
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This document provides an overview of congenital infections caused by cytomegalovirus (CMV) and Toxoplasma gondii. It discusses the properties and transmission of CMV, which can cause birth defects if transmitted from mother to fetus. Clinical manifestations range from asymptomatic infection to organ damage in immunocompromised individuals. Diagnosis involves virus detection and antibody testing. Treatment focuses on antiviral drugs for severe cases. Toxoplasmosis is caused by the parasite T. gondii and transmitted through cats, meat, and congenitally. It can cause abortion or symptoms in susceptible hosts. Diagnosis is usually serological and treatment includes sulfa drugs and pyrimethamine.
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Ventilator-associated pneumonia (VAP) is a type of hospital-acquired pneumonia that occurs in patients on mechanical ventilation for more than 48 hours. It is diagnosed using clinical criteria like fever, leukocytosis, and radiographic evidence of pneumonia combined with microbiological testing of respiratory samples. Treatment involves administering antibiotics based on the local hospital antibiogram, with empiric therapy targeting likely gram-positive and gram-negative pathogens. Prevention strategies focus on reducing ventilator days through daily weaning assessments and using bundles of care involving oral hygiene, elevation of the head, and peptic ulcer/DVT prophylaxis.
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This document summarizes various adult immunization guidelines. It discusses vaccines for tetanus, diphtheria, pertussis, human papillomavirus, influenza, varicella, herpes zoster, measles, mumps, rubella, pneumococcus, meningococcus, hepatitis A, hepatitis B, and Japanese encephalitis. For each vaccine, it provides information on disease burden, target populations for vaccination, vaccine types, dosing schedules, efficacy, contraindications and precautions. The document was written by Dr. Sshrutkirti Gupta and provides a comprehensive overview of recommended adult immunizations.
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2) Types of media such as solid, liquid, enriched, selective, differential and special media formulated for specific purposes.
3) Culture methods like streaking, lawning, stabbing and pour plating used to isolate bacteria and study their properties.
4) Requirements for culturing anaerobic bacteria in specialized conditions with low oxygen levels.
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This document discusses autoimmunity and autoimmune diseases. It defines autoimmunity as the immune system mistakenly attacking the body's healthy tissues, outlines several potential causes, and notes that over 80 diseases can result. It then covers mechanisms of autoimmunity, classification of diseases as organ-specific or systemic, associations with HLA types, diagnostic criteria and methods, examples of diseases, and challenges with treatment.
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Academy of Oriental Medicine at Austin
Academy of Oriental Medicine at Austin
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About AOMA: The Academy of Oriental Medicine at Austin offers a masters-level graduate program in acupuncture and Oriental medicine, preparing its students for careers as skilled, professional practitioners. AOMA is known for its internationally recognized faculty, award-winning student clinical internship program, and herbal medicine program. Since its founding in 1993, AOMA has grown rapidly in size and reputation, drawing students from around the nation and faculty from around the world. AOMA also conducts more than 20,000 patient visits annually in its student and professional clinics. AOMA collaborates with Western healthcare institutions including the Seton Family of Hospitals, and gives back to the community through partnerships with nonprofit organizations and by providing free and reduced price treatments to people who cannot afford them. The Academy of Oriental Medicine at Austin is located at 2700 West Anderson Lane. AOMA also serves patients and retail customers at its south Austin location, 4701 West Gate Blvd. For more information see www.aoma.edu or call 512-492-303434.
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Lab diagnosis-viruses.ppt
- 1. Laboratory Diagnosis of Viral Infection Dr. Sshrutkirti Gupta
- 2. • Collection of samples • Transport • Cultivation • Methods of diagnosis Following Points will discussed today
- 3. Importance of diagnosis of viral infection • Rubella diagnosed in 1st trimester-abortion is recommended. • If baby born to HBV Surface Ag positive mother, immunization at birth is mandatory. • Diagnosis of diseases caused by viruses for which antiviral treatment is available (eg. Herpes) • Screening of HIV ,HBV helps in preventing transmission of the disease • Early detection of epidemics like influenza, helps to initiate appropriate control measures.
- 4. Sample collection • Respiratory tract- Nasal washing, throat swab, nasal swab, sputum (adeno, influenza, enterovirus, orthromyxo, paramyxo, HSV, Rubella) • Gastrointestinal tract-stool, rectal swab (Reovirus, Rotavirus, Adenovirus, norwalk calcivirus) • Maculopapular Rash-Throat swab, rectal swab (Adeno, entero, Rubella, measles)
- 5. • Vesicular rash-Vesicle fluid, scraping or swab Coxsackie echo, HSV, VZV • Central Nervous system- Picorna- Stool , Arbovirus- rarely cultured Rabies- Tissue, saliva, brain biopsy HSV, CMV, Mumps, Measles- CSF, Biopsy • Urinary Tract Infection- Adenovirus, CMV-urine • Blood born infections HIV,HTLV,HBV,HCV,HDV-Blood ….Cont Sample collection
- 6. Methods of diagnosis • Direct microscopy- virus • inclusion bodies • Isolation of virus/cultivation • Antigen detection • Antibody detection • Nucleic acid detection
- 7. Microscopy • A-Electron microscopy • B- Fluorescent microscopy • C- Light microscopy Electron Microscopy • Demonstration of viral particles in clinical samples • Mainly for those whose cultivation is difficult • Immunoelectron microscopy – more sensitivity Rabies virus
- 9. Direct demonstration-EM Sample • Faeces • Vesicular fluid • CSF • Urine Virus Rotavirus, HAV, Adeno, Norwalk, astrovirus. Herpes simplex, VZ Enterovirus, VZ CMV
- 10. EM structure of Flu virus
- 11. Microscopy Fluorescent microscopy Rabies Paramyxovirus, Orthomyxoviruses, Adenovirus Herpesvirus
- 13. Light microscopy Inclusion bodies • Rabies- negri bodies • Vaccinia (Guarnieri bodies) • Molluscum contagiosum (Molluscum bodies) • Cowdry TypeA- Herpes, Yellow fever • Cowdry type B-Adeno, Polio Stains- Giemsa, Eosin-methylene blue, Haematoxylin
- 16. Cultivation • By inoculation in to animals • By inoculation into embryonated egg • By inoculation into cell culture • Sample process to remove bacterial contamination
- 17. Animal inoculation • Suckling mice- Arbovirus, Coxsackie • Different routes- Intracerebral, subcutaneous, intraperitoneal, intranasal • Observe for signs of disease, death • Tissue tested for presence of virus- neutralisation test (antisera), demonstrate inclusion bodies
- 19. Egg Innoculation Site • Chorioallantoic Membrane • Allantoic cavity • Amniotic cavity • Yolk sac Virus • Pox (pocks)-variola or vaccinia • Influenza, yellow fever • Influenza • Chlamydiae, rickettsiae
- 20. Tissue culture • Organ culture • Explant culture- rarely used • Cell culture
- 21. Organ culture • Bits of organ maintained in tissue culture growth medium • Used mainly for highly specified parasites of certain organ eg- corona virus- Tracheal ring culture
- 22. Cell Culture • Primary cell culture • Diploid cell culture • Continuous cell line
- 23. Primary cell culture Limited growth up to 5-50 divisions Use- primary isolation, vaccine production Examples of cell lines 1. Rhesus monkey kidney cell culture 2. Human amnion cell culture 3.Chick embryo fibroblast cell culture
- 24. Diploid cell strain • cells of single type –contains same no. of chromosomes as in the parent cell • Limited growth 50 cell division • Developed from fibroblast • WI-38-human embryonic lung cell strain Rabies virus vaccine HL-8 (Rhesus embryo cell strain)
- 25. …Cont Cell culture Continuous cell line • Single type capable of unlimited growth eg- 1. HeLa (Human carcinoma of cervix cell line) 2. HEP-2(human epithelioma of larynx cell line) 3. Mckoy (human synovial carcinoma cell line) 4. BHK-21 ( Baby hamster kidney cell line)
- 26. Detection of growth in cell culture • 1 Cytopathic effect • 2 Haemadsorption • 3 Interference • 4 Transformation • 5 Immunoflurescence • 6 Electron microscopy • 7 Detection of enzymes
- 27. Cytopathic effects a.Syncytium formation- measles, RSV neighbouring cell fused together to form multinucleated giant cell. b. Cell necrosis & lysis- Enterovirus c. Roundening of cells- Picornavirus viral replication leads to neclear pyknosis , rounding, refratility, degeneration d. Discrete focal degneration- Herpes
- 29. Herpes
- 30. Cell necrosis by enterovirus
- 31. Rounding of cells-polio virus
- 32. Rounding of cells- poliovirus
- 36. …Cont Cytopathic effects • Haemadsorption- Influenza, parainfluenza,Toga –express Haemagglutinin on infected cells • RBC suspension layered on cell layer • RBC adsorb on infected cells
- 37. Antigen detection • Immunoflurescence • Serological tests Antibody detection • Serological tests
- 39. Newer techniques • Molecular • DNA PCR • RNA PCR