Virology 2024 | Microbes with Morgan 2024

1. Margie Morgan, PhD,
MT(ASCP), D(ABMM)
1

2.  Specimens suspect for routine viral pathogens can be processed in a
laboratory using BSL-2 precautions. These would be viral pathogens that will
not generally cause fatal disease, such as Herpes simplex virus.
 BSL-2 precautions include processing specimens using a BSL2 HEPA–
filtered biosafety cabinet, wearing a front closed laboratory gown, and gloves
2

3. 1. Direct Staining Cells for Antigen
2. Enzyme Immunoassay
3. Viral Cell Culture
4. Molecular Amplification
3

4.  Direct Fluorescent antibody (DFA) stain for HSV1/2 & VZV
 Collect cells from base of fresh vesicular lesion
 Stain with antibody specific for HSV 1/2 and/or VZV
 Look for fluorescent cells (virus infected) using fluorescence microscope
 Sensitivity @ 80% compared to molecular assays
 Tzanck prep
Cells from lesion stained with Giemsa stain /examine for
multinucleated giant cells consistent with Herpes virus family
Sensitivity @ 50% / Cannot differentiate HSV 1, HSV 2, or VZV
Tzanck DFA
4

5. • Enzyme immunoassay (EIA) –
 Antigen in specimen forms a complex with test antibody, then a color producing
substrate is added and binds to Ag/Aby complex
 Used for rapid point of care testing (20 min), moderate sensitivity (70%
compared to molecular assays) positive with high viral load, specificity adequate
during high prevalence viral season
 Used most often for respiratory virus testing:
 Influenza A /B, Respiratory syncytial virus (RSV), SARS-CoV-2
• Membrane lateral flow EIA Liquid/in-well EIA
5

6. Inner wall of container coated with monolayer of cells
grown in enriched liquid growth media
• Three types of cell lines used for culture:
 Primary – obtained directly from animal or human organ, processed
and placed into culture vial, innate animal viruses can make culture
reading difficult
 Example: Rhesus monkey kidney-RMK
 Diploid – semi continuous cell lines– Can survive 20 – 50
subcultures
 Example: Human diploid fibroblast cells (MRC-5-Microbiology
Research Council 5)
 Continuous cell lines – survive continuous passage (immortal)
 Example: Tumor lineages such as HEp-2 and HeLa
6

7.  Patient specimen inoculated onto cell monolayer, incubated
for a specified number of days, then read under light
microscopy for “Cytopathic effect” – the effect viral growth
has on the cells
• The pattern of destruction of the cell monolayer is specific
for each virus type and the time in which destruction
occurs is virus specific.
• Sensitive but slow <= 21 days
 Research and study of viruses
7

8. Spin Down Shell Vial Culture
• Technique created to speed up viral cell culture
• Cell monolayer is prepared on a coverslip
• Specimen inoculated into vial with coverslip and growth
medium
• Centrifuge vial to rapidly induce virus invasion of cells
• Incubate @ 35*C for 24-72 hours
• Perform DFA stain to search for early virus antigens
Cover slip
8

9.  Molecular Amplification of DNA or RNA (Qualitative)
• DNA viruses - direct amplification of genetic material
• RNA viruses (RT-PCR) – reverse transcriptase enzyme transcribes
complimentary copies of DNA using the RNA template
• More rapid and may exceed sensitivity of viral culture. Is the current
Standard of Practice for detection of most virus types:
 Respiratory, encephalitis, lesion, and diarrhea…….
• Test of diagnosis not cure – can shed residual virus for 7 – 30
days after initial positive test for acute infection
 Quantitative molecular assays are used for assessing quantity of
virus in the specimen to help access severity of infection: used with
CMV, Hepatitis B and C, and HIV
9

10.  Viral transport media (VTM) or Universal transport media
(UTM) is a Hanks balanced salt solution, protects DNA &
RNA, antibiotics to prevent bacterial overgrowth during
transportation
• A must for swab collected specimens: lesions, mucous membranes,
nasopharyngeal & throat
• Used for cell culture and molecular amplification assays
 Storage of specimens prior to testing:
• Short term media storage at 4˚C
• Long term (>72hours) media storage at -80˚C
 VTM/UTM is filtered (45nm filter) to eliminate
contaminating bacteria prior to cell culture inoculation
10

11. 11

12.  Enveloped double stranded DNA virus
 Eight human Herpes viruses
• Herpes simplex 1
• Herpes simplex 2
• Varicella Zoster
• Epstein Barr
• Cytomegalovirus
• Human Herpes 6, 7, and 8
 Latency (hallmark of Herpes viruses) occurs
within low numbers of specific kinds of cells
on human tissue, the cell type is different
for each Herpes virus
12

13.  Transmission: direct contact/secretions
 HSV replicate in epithelial cells producing vesicles, the virus then
ascends sensory nerves to the dorsal root ganglia. There is initial
replication then latency in dorsal root ganglia.
 Reactivation infection spreads from the ganglia.
 Infections:
• Gingivostomatitis - common primary infection
• Herpes labialis
• Ocular
• Encephalitis
• Disseminated disease in immune suppressed
• Neonatal – If suspicious due to mother possibly being HSV positive, skin
surface viral surveillance PCR assays should be performed on the newborn
Therapy – Acyclovir and Valacyclovir
13

14.  Viral Cell Culture
• Produce CPE within 24-48 hrs
• Human diploid fibroblast cells (MRC-5)
• Produces characteristic CPE
Negative
fibroblast
Cell line
HSV CPE consists of cell
rounding starting on the
edge of the monolayer.
Histology/Cytology – Observe for
multinucleated giant cells, cannot
differentiate HSV 1, 2, and VZV
Molecular Amplification -Standard
of practice for detecting HSV 1/2 from
patient specimens including CSF
Serology – used to screen for past
infection, not for acute diagnosis
14

15.  Transmission: close contact
 Replication in epithelial cells ascend to the dorsal root
ganglia to establish latency.
 Diseases:
• Chickenpox (varicella) primary infection
 Considered a disease of childhood, adults and
immune suppressed more severe symptoms with
possible progression to pneumonia and/or
encephalitis
• Shingles (zoster) recurrent infection
• Lesions usually appear over one isolated
dermatome and capable of dissemination
• Ramsay-Hunt syndrome – facial nerve infected
leads to facial paralysis (pictured)
15

16.  Histology – Multi-nucleated giant cells
 Serology – 1º used for immune status check
 Cell culture – growth in 5-7 days, fibroblasts
Sandpaper like appearance in cell monolayer
with scattering of rounded cells
 Molecular amplification (PCR) for disease diagnosis
is standard of practice
 Preventative vaccine for children and adults
16

17.  Transmitted by blood transfusion , vertical and horizontal
transmission to fetus, close human contact, and sexual contact
 Latency: Macrophages
 Infection:
• Initial infection asymptomatic in most and usually occurs in the first
decade of life
• Congenital infection– most common cause of TORCH infection
• Perinatal acquisition of infection can occur
17

18.  Mononucleosis – fever, absence of swollen lymph nodes, and
heterophile antibody test is negative (differs from EBV
mononucleosis)
 Organ specific disease can occur, primarily in the immune
suppressed host
• Gastrointestinal
• Hepatic
• Neurological
• Cardiovascular
• Primary infection is more serious than recurrent disease
18

19.  Laboratory Diagnosis:
• Cell culture on Human diploid fibroblast cells
 Cytopathic effect can take >=14 days
 CPE described as grape like clustering of rounded cells
• Quantitative PCR is best
 Due to persistent CMV shedding from past infection
quantitative PCR used to detect significant viral load
associated with both primary or recurrent infection
• Histopathology – Infects epithelial cells,
macrophages and T lymphocytes. Intranuclear
and intracytoplasmic inclusions classic (Owl eye)
 Treatment: Ganciclovir, Foscarnet, and Cidofovir
19

20.  Transmission: close contact and saliva
 CD21 cell receptor for B lymphocyte cell invasion and
proliferation
 Latency in the B lymphocyte
 Diseases include:
• Infectious mononucleosis
 Diagnosis: MonoSpot test – specific heterophile antibody
produced in infection, patient serum agglutinates with heterophile
horse red blood cells in reaction
• Lymphoreticular disorders
• Oral hairy leukoplakia
• Burkitt’s lymphoma
• Nasopharyngeal Carcinoma
• 1/3 of Hodgkin’s lymphoma cases
 Diagnosis: Will not grow in viral cell culture
• Serology and molecular assays (PCR) provide diagnosis
Examination of peripheral
WBC’s in EBV infection
can demonstrate enlarged
reactive T lymphocyte
20

21. VCA IgM VCA IgG EBNA-1 IgG
Negative Negative Negative No immunity
Positive Negative Negative Acute infection
Positive Positive Negative Acute infection
Negative Positive Positive Past infection
Negative Positive Negative Acute or past infection
Positive Positive Positive Late primary infection
Negative Negative Positive Past infection
VCA = viral capsid antibody
EBNA = EBV nuclear antigen
Serologic Diagnosis of EBV
Past
infection
21

22. HHV-6
 Roseola [sixth disease] usual age 6m-2yr, high
fever and rash
 Adult – encephalitis in immune compromised
 Note: HHV-6 undergoes germ line integration
during childhood infection and can be a
source of false positive results in the normal
adult host when performing a CSF PCR, must
be cautious in test result interpretation
HHV-8
 Kaposi’s sarcoma - Lesions in soft tissues. the
skin, lymph nodes, internal organs, and mucous
membranes. It often infects patients with immune
deficiencies, such as HIV/AIDS.
22

23.  Castleman disease
 Rare disorder that involves an overgrowth of cells in lymph
nodes and related tissue (lymphoproliferative disorder) There
are different forms with different findings and prognosis.
 Primary effusion lymphoma
• Most often in HIV/AIDS patients
• Localized in body cavities without a tumor mass.
• Large cells , large round to irregular
nuclei and prominent nucleoli
Classic Lollipop
like lesion
23

24. Adenovirus
24

25.  DNA - non enveloped/ icosahedral virus
 Latency: lymphoid tissue
 Transmission: respiratory and fecal-oral route
 Diseases:
• Pharyngitis (year-round epidemics)
• Pneumonia
• Gastroenteritis in children
 Adenovirus types 40 & 41 most common
• Kerato-conjunctivitis – usually bilateral red
painful and inflamed eyes for @ 2 weeks
• Disseminated infection in immunosuppressed –
usually pneumonia, @ 75% fatality/ no therapy
• Hemorrhagic cystitis in immune suppressed /
particularly in transplantation
25

26. • Cell culture (CPE), <=5 days
• Molecular methods (PCR) superior
• Stool (Adeno strains 40/41) PCR & Ag tests
• No specific viral therapy
• Histology - Intranuclear inclusions known as
smudge cells Basophilic and nuclear membranes
become blurred
Cell culture: Round cells
2 – 5 days with stranding
– HeLA or Hep-2 cells
26

27. Parvovirus
Canine parvovirus is a contagious virus mainly
affecting dogs. CPV is highly contagious and is
spread from dog to dog by direct or indirect
contact with their feces. Vaccines can prevent
this infection, but mortality can reach 91% in
untreated cases. This is not contagious to
humans.
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28.  Non-enveloped ss DNA virus infects humans
 Infections:
• Erythema infectiosum (Fifth disease) – childhood
infection with headache, rash on cheeks, and cold-like
symptoms
• Hydrops fetalis – infection in pregnant, can lead to
miscarriage
• Chronic hemolytic anemia in patients with HIV/AIDS
• Arthritis and Arthralgia
• 20% of infected do not have symptoms
 Histology – In bone marrow, virus infects mitotically active
erythroid precursor cells
 Molecular and Serologic methods
confirm histology to establish diagnosis
Slapped face appearance
of fifth disease
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29. Papillomavirus
Polyomavirus
Non-enveloped double stranded DNA viruses
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30.  Skin and anogenital warts
 Benign head and neck tumors
 Cervical and anal intraepithelial neoplasia and cancers
 High risk HPV types 16, 18 cause @ 70% of cervical cancers.
 Other high-risk types include: 31, 33, 45, 52, 58
 Low risk HPV types 6 and 11 cause @ 90% genital warts
 Diagnosis :
 Pap Smear
 Molecular methods: Detection of HPV and typing of HPV types
 Guidelines suggest both PAP and molecular HPV testing for women 30 - 65
years of age / performed every 5 years
 Vaccines - HPV 6,11,16,18,31,33,45, 52 and 58 for young females and males
Abnormal Pap smear
30

31. • JC virus
 Progressive multifocal leukoencephalopathy (PML)
 Encephalitis in AIDS, cancer and immune suppression
 Demyelination seen in MRI scans
 Diagnosis: CSF specimen / PCR assay
 Histology: Destruction of oligodendrocytes in brain with
formation of giant glial cells
• BK virus
 Virus maintains latency in kidney causing nephropathy and a
hemorrhagic cystitis
 Activates and progresses with immune suppression
 Diagnosis: Quantitative PCR (plasma)
 Histology: Homogenous and purple intranuclear
inclusions, primarily in tubular epithelium
Giant Glial Cells of JCV
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32. Hepatitis B
Ground glass hepatocytes of chronic Hepatitis B
32

33.  Enveloped/ double stranded DNA (an envelope aids survival to pass
between cells, but can be inactivated with detergents)
 Acquisition from Hep B positive mother during birthing process
 Spread by contact with blood and other body fluids
 Spectrum of Hepatitis B symptomatology
• Acute phase - subclinical hepatitis to icteric hepatitis fulminant
• Chronic phase - chronic hepatitis, cirrhosis, and hepatocellular carcinoma
 Vaccinate to prevent
 Therapies effective for patients with minimal liver damage
 Diagnosis
• Serology and Molecular assays to detect virus and viral load for
severity of infection
33

34.  Surface Antigen Positive
• Patient has Active Hepatitis B or is a Chronic Carrier
 Next perform Hep B Quantitation to assess viral load and
 Perform Hep e antigen test – if positive, patient is a chronic carrier
of HBV. This carries a worse prognosis
 Core Antibody Positive
• Immune due to prior infection,
acute infection or chronic carrier
Surface Antibody Positive
• Immune due to prior infection
or vaccine
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35. Hepatitis C
Mosquito borne Flaviviridae:
Dengue
Zika
Yellow fever
West Nile
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36.  Single stranded RNA virus
 Disease acquisition:
Parenteral transmission, drug abuse, sexual, blood products or organ
transplants (prior to 1992), poorly sterilized medical equipment
 Infection: humans and chimpanzees
 Chronic Hep C common in US with estimated 3.2 mil
• Acute self-limited disease that progresses to a disease that mainly
affects the liver
• Seven genotypes (1-7), Type 1 virus most common in USA
• Infection persists in @ 75-85% without symptoms
• 5 - 20 % develop liver cirrhosis
• 1-5 % develop hepatocellular carcinoma
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37. Diagnosis:
• Hepatitis C IgG antibody detected
• Must then perform:
 RNA quantitative assay to assess viral load
 FibroScan to assess liver damage/ therapy
cannot reverse cirrhosis
 Genotype of virus determined and aids in
proper therapy selection and duration
 No vaccine available
 Antivirals available capable of cure >= 85% of
patients Periportal mononuclear
inflammatory infiltrates and
mild interface activity.
37

38. • Dengue – “break bone fever”
• Vector: Aedes aegypti mosquito /Endemic - Africa, the Americas,
the Eastern Mediterranean, South-East Asia and Western Pacific
• Disease
 Fever, rash, and severe joint pain
 Small % progress to a hemorrhagic fever which can be fatal
 Diagnosis: Serology / IgM for acute infection/ Molecular assays (serum)
 Zika virus
• Vector: Aedes aegypti and A. albopictus mosquitoes
• Outbreak 2016 in Brazil with spread to central America, Caribbean and US (Miami)
• Milder primary disease than Dengue in adults, with only fever and rash
• Neurologic tropism makes it very problematic during pregnancy
 Microcephaly in fetuses borne to infected moms
 Developmental issues can occur in young children infected with ZIKA
 Guillain-Barre syndrome post infection sequelae in adults
• Diagnosis: Serum IgM / Molecular assays serum, urine, amniotic fluid and CSF
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39. Yellow fever
 Vector – Aedes aegypti mosquito
 Outbreak Brazil 2018 /large numbers of infected mosquitoes thrived in
heavily populated Brazilian cities
 Endemic in Africa, Central and South America
 Most cases mild with 3-4 days fever, headache, chills, back pain,
fatigue, nausea, vomiting
• 15% experience liver damage (jaundice) and hemorrhagic issues (20 – 50%
fatality rate)
 Diagnosis:
• Molecular testing for virus in serum
 No specific anti-viral therapy
 Vaccine – supplies life-long immunity
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40. • West Nile
 Vector: Culex mosquito
 Common across the US over the lasts two decades
 Bird primary reservoir, humans and horses at risk
 Disease
 80% asymptomatic
 20% fever, headache, muscle weakness
 Small % progress to encephalitis, meningitis, flaccid paralysis
 Serology and Molecular assays available for diagnosis
 Molecular assays for CSF have low sensitivity due to narrow window of time when virus is
present, both serology (IgM) and molecular methods are needed for diagnosis
 A positive WNV IgM with a clinical history and epidemiology usually diagnostic
NOTE: Cross reactions occur in serologic testing within the Flavivirus family:
Dengue, Zika and West Nile virus can be diagnostically confused/
Molecular assays and neutralization testing can provide specific results 40

41.  Chikungunya virus
 Vector: Aedes mosquito including aegypti
 Origin in Asia and African continents with recent migration to the
Caribbean and SE USA
 Acute febrile illness with rash followed by joint pain, both disease and
location similar to Dengue, but no hemorrhagic phase
 When screening a patient for Chikungunya,
 May need to rule out infection with Dengue and Zika,
 Similar symptoms but very different sequelae.
 Specific molecular assays can assist with diagnosis.
41

42.  >20 outbreaks since discovery in 1970’s/ ss RNA virus
• Animal reservoir : fruit bat
• Outbreaks in Uganda (2022) Sudan strain and West Africa
(DRC, Guinea) Zaire strain (2013)
 Human becomes contagious when symptoms develop which include,
fever, weakness, myalgia, and headache, as disease moves on there
can be a “wet” phase with copious vomiting and diarrhea and
sometimes a hemorrhagic phase with bleeding
• Patients are transferred to special pathogen containment units to handle
unique patient care biohazardous risks and waste disposal needs
• Biothreat Level A pathogen, high level PPE and containment
• Multifocal necrosis in liver, spleen, kidneys, testes and ovaries
• Fatality rate >=55%
 Public health and special pathogen unit molecular testing, with the virus
detectable at >= 4 days of illness
Filoviridae:
42

43. Severe Acute Respiratory Syndrome 1 (SARS)
o Outbreak in China 2003 that spread to 29 countries / @ 80 cases in the US
o Bat coronavirus with civet cat identified as the intermediate host
o Dry cough and/or shortness of breath with pneumonia by day 7-10 of illness
o Laboratory testing available only in public health laboratories (CDC) Molecular
tests performed on NP, Throat, sputum, blood, and stool specimens.
Most coronaviruses cause cold like symptoms,
but a few took a very different path.
Middle East Respiratory Syndrome (MERS)
o Endemic in Arabian peninsula (1st reported 2012)
o Infection from direct contact with infected camels
o Close human to human contact may transmit infection but no outbreaks
o Fever, rhinorrhea, cough, malaise followed by shortness of breath
o 30% fatality rate from respiratory failure
o
43

44. • Bat coronavirus transmitted, ? intermediate host, possibly Raccoon dog
• Human spread virus primarily by respiratory droplet
• Influenza-like illness ranging from asymptomatic and mild symptoms to severe respiratory
illness with systemic complications & long-term complications
 Most common presenting symptoms: Fever, dry cough, shortness of breath, sore throat,
nasal congestion, loss of sense of smell and taste, and diarrhea
 Transmission, symptoms and severity can be COVID variant dependent and patient
specific, worse for obese, cardiac issues, diabetes, asthma, and immune suppression
.
• Diagnosis:
 Optimal specimen: Nasopharyngeal collected in universal transport medium (UTM)
 Detection of viral RNA by molecular amplification assay most sensitive and specific
• Antibody detection (IgG)to detect past COVID infection or response to vaccine
• Vaccine: protection, certainly provides protection against serious infection
44

45. 45

46.  Diverse group of > 60 viruses – SS RNA
 Seasonal viruses most often in summer and fall
• Polio virus/poliomyelitis, CNS infection leads to flaccid paralysis
 Salk Inactive Polio Vaccine (IPV) recommended
• Coxsackie A = vesicular oral lesion (Herpangina)
Hand, foot, and mouth disease
• Coxsackie B = Pericarditis and Myocarditis
• Enterovirus, emergence of two pathogens
 (1) Parechovirus - aseptic meningitis in children, fever and rash
 (2) EV-D68 meningitis with acute flaccid myelitis
• Echovirus = various infections, intestine
• Rhinoviruses = common cold
 Grow in continuous cell lines in 5-7 days
• Teardrop or kite like CPE formed
 Molecular assays gold standard for diagnosis
46

47.  Fecal – oral transmission, contaminated food or person to person
 Common traveler’s disease
 Recent outbreaks in US homeless populations due to inadequate
sanitary facilities
 Incubation 15 – 50 days with abrupt onset, low mortality, no carrier
state
 80% infected develop symptoms of jaundice & elevated
aminotransferases
 Diagnosis – serology with diagnostic IgM positive in early infection
 Antibody (IgG) is protective and lasts for lifetime
 Vaccination to prevent
47

48.  Single stranded RNA virus
 Hemagglutinin (H) and Neuraminidase (N) spikes on outside of the
viral capsid
• H and N provide virus strain typing information - H1N1, H3N2
 Antigenic drift - minor change in the amino acids of either the H or
N glycoprotein, can occur yearly
 Cross antibody protection exists that provides some protection
 Antigenic shift - genome reassortment with a “new” virus
created/origin usually bird or animal (potential pandemic)
 H5N1 = Avian Influenza strain
 H1N1 = 2009 Influenza A pandemic strain
48

49.  Disease: Acute onset of respiratory symptoms (nose, throat or lung) which can
progress to secondary bacterial lung infection
 Influenza season usually @ November through February
 Usually, one H and N type dominates each season most recently H3N2 and H1N1
 Diagnosis
• Molecular assays best for influenza diagnosis
 Tests detect both Flu A and the H/N type, non-typable Flu A could be unique virus
• Antigen tests are rapid but lack sensitivity (@75%)
 Treatment: Tamiflu (Oseltamivir)
• Tamiflu has remained sensitive to evolving strains
 Influenza B
• Milder form of respiratory symptoms compared to Flu A
• Usually <=10% of cases of total influenza cases during influenza season
 Vaccinate – Quadrivalent vaccine that contains 2 A types and 2 B types
49

50.  Measles (Rubeola)
• Fever, rash, coryza, cough and conjunctivitis. Very
contagious
• Koplik’s spot – small red spot with central bluish
discoloration- seen in the inner lining of the cheek
 Subacute sclerosing panencephalitis
 Rare form of chronic progressive brain inflammation
caused by slow infection with certain defective strains of
hypermutated measles virus.
 Diagnosis: Clinical symptoms plus molecular tests
performed on nasal or throat specimens
 Histology for acute lung injury – multinucleated
giant inclusions with perinuclear halos
 Vaccinate – MMR (Measles, Mumps, Rubella)
vaccine
 Treatment: Not specific, supportive
H and E stain/ lung
50

51.  Types 1, 2, 3, and 4
 Spread by respiratory droplets
 Disease:
• Can occur throughout the year
• Upper respiratory tract infection in adults and children with
fever, runny nose and cough
• Lower respiratory tract infection - Croup, bronchiolitis and
pneumonia more likely in children, elderly and immune
suppressed
 Molecular assays for diagnosis
 Supportive therapy only
 No vaccine
51

52.  Spread by respiratory droplets and contact
 Infection of parotid and salivary glands, other
sites affected less commonly: testes/ovaries,
Eye, Inner ear, CNS
 Diagnosis: clinical symptoms, serologic tests,
and molecular assays
 Histology – diffuse interstitial edema and an
inflammatory infiltrate composed of histiocytes,
lymphocytes, and plasma cells.
 Vaccination: Measles/Mumps/Rubella (MMR)
 No specific therapy, supportive
Mumps Orchitis
52

53.  Respiratory disease - common cold to pneumonia, bronchiolitis to croup
(young infant) more serious in infants and immune suppressed
 Transmission by contact and respiratory droplet
 Best specimen for testing: Nasopharyngeal or nasal aspirate
 Diagnosis: Molecular assays
 Lung biopsy: Syncytium formation in tissue
 Treatment: Ribavirin Vaccinate for prevention
Classic CPE =
Syncytium
formation(multinucleated
giant cell formation) in
HeLa cell line
Syncytium formation
In lung tissue
53

54.  1st discovered in 2001 – seasonal (winter) community
acquired viral respiratory tract disease
• 95% of cases in children <6 years of age but can be seen
in the elderly and immune suppressed
• Upper and lower respiratory tract disease
 2nd only to RSV in the cause of bronchiolitis
 Will not grow in viral cell culture
 Molecular assays for diagnosis
 Treatment: Supportive, no specific anti-viral therapy
available
 No vaccine
54

55.  Nonenveloped/double stranded RNA virus
 Winter - spring seasonality
• Gastroenteritis with vomiting – common cause of severe
diarrhea in children 6m – 2 yr, can lead to dehydration
• Fecal – oral spread and can be transmitted by aerosols
• Resistant to detergents / hardy non-enveloped virus
 Major cause of childhood death in developing countries
 Diagnosis – unable to grow in cell culture
• Molecular assays of stool for diagnosis
• Vaccine (oral) has greatly decreased infection rate
Rota = Wheel
EM Pix
55

56.  Non enveloped single stranded RNA virus
 Spread by contaminated food and water, feces and vomitus, direct
contact with sick, and touching contaminated surfaces, unfortunate
association with cruises
 <=20 virus particles in waste material can transmit infection making
Norovirus infection very contagious
 Leading cause of epidemic gastroenteritis in all age groups, particular
problem in the elderly
• Vomiting and diarrhea
• Disease course usually lasts 24-48 hours
• No specific therapy / No vaccine
 Diagnosis: Molecular assays /stool specimen
56

57.  Unique characteristics of HIV
• CD4 primary receptor site on lymphocyte for attachment and entry of
Retrovirus
• Reverse transcriptase enzyme converts genomic RNA into DNA
 Transmission - sexual, blood and blood product exposure,
perinatal
 Non-infectious complications of HIV/AIDS:
• Lymphoma, Kaposi’s sarcoma, anal cell carcinoma, and non-
Hodgkin’s lymphoma
 Infectious complications:
• Pneumocystis, Cryptococcal meningitis, TB and Mycobacterium
avium complex, Microsporidia, Cryptosporidium, STD’s , Hepatitis B
and C
57

58. Old way to test -Antibody Enzyme immunoassay with Western Blot confirmation
 Positive serology (EIA) test confirmed by Western blot performance
 Western blot detects gp160/gp120 (envelope proteins), p 24 (core), and
p41(reverse transcriptase)
 Must have positive serology assay plus at least 2 solid bands on Western blot to
confirm a positive result for HIV diagnosis – positive detected at @ 3 months
Improved testing - Antigen/antibody combination (4th generation) immunoassay that
detects IgG and IgM HIV-1/HIV-2 antibodies plus HIV-1 p24 antigen.
Detects HIV infection earlier (at 18-45 days of infection) than previous method and
screens for both acute and established infection.
Patients testing positive are immediately placed on therapy and need additional testing:
 HIV viral load quantitation using quantitative PCR methods
 Resistance gene testing – viral subtype knowledge used to optimize therapy
 Most isolates in US / type B
 Monitor for low CD4 counts for HIV infection severity
58

59. RNA Virus
Rubella
59

60.  Rubella, Three-day measles, or German measles
Relatively mild disease usually in children presenting with rash, low grade fever,
cervical lymphadenopathy
Respiratory transmission
 Congenital rubella –
• Occurs in a developing fetus of a pregnant women who has contracted Rubella
• It was the neurotropic virus of the fetus prior to the occurrence of Zika
 Infection caused deafness, eye abnormalities, congenital heart disease in the
newborn
 Diagnosis: serology in combination with clinical symptoms
 Live attenuated vaccine (MMR) to prevent
60
Rash

61. Hantavirus
61

62.  1st outbreak occurred in 1993 on a farm in the four corner states
(NM,AZ,CO,UT), this led to disease description
 2012 outbreak in Yosemite National Park where mice worked their way into
insulation in the walls of cabins for rent
 Most common in animals west of Mississippi River
 Transmission
• Urine and secretions of wild deer mouse and cotton rat
• Transmission from pet rats to humans
 Respiratory disease that can lead to respiratory failure, symptoms usually
begin with myalgia, headache, cough
 Diagnosis: Serology
 Supportive therapy
62

63. Smallpox virus Variola virus
Vaccinia virus
Monkeypox
Molluscum contagiosum
63

64.  Variola virus – Smallpox, eradicated in 1977
 Vaccinia virus - Smallpox vaccine strain,
immunologically related to smallpox
• Vaccinia can also cause limited disease after vaccine
administration in immune suppressed which would prevent
vaccination of this population
 Smallpox begins as maculopapular rash on central
body moving outward and progressing to a vesicular
rash
 BSL4 conditions are required for laboratory work on
Variola virus / Category A Biothreat agent
 Possible infection reported to public health for case
investigation and molecular diagnostic testing
64

65.  Historically, related to exposure to infected exotic
animals, endemic to Africa (DRC and Nigeria)
 Worldwide human Mpox outbreak, began May 2022 in
Europe
 Symptoms: fatigue, fever, headache, aches and pains.
Over the next week, Characteristic deep seated, well circumscribed
umbilicated lesions develop (pictured)
 Transmission by close contact
 Diagnosis: Detect virus from lesions using molecular
assay
 Antiviral therapy (Tecovirimat)
 Vaccination can assist with prevention
65

66.  Appears as small flesh-colored papules on the skin
 Commonly involved areas include groin and axillae but
 disseminated disease may be seen in immune
compromised
 Treatment dependent upon patient, location and extent of
lesions
 Histopathology shows a well
circumcised epithelial
proliferation with central
depression
66

67. Rabies virus
67

68.  Worldwide in animal populations
• Bat and raccoon primary reservoirs in US
• Ferrel dog reservoir in developing countries
 Infections from bites and inhalation of animal aerosolized saliva, urine
and feces
 Post exposure rabies vaccine and rabies immunoglobulin PRIOR to
the development of symptoms prevent disease development
 Classic disease symptom: excessive salivation due to paralysis of
throat muscles
 Diagnosis: Detection of viral particles (Negri bodies) in the brain and
detection of rabies genetic material testing saliva by molecular
assays
 Public health laboratories for assistance with diagnosis and therapy
Bullet shaped viral
particles
68
Intracytoplasmic Negri
bodies in brain biopsy

69.  Creutzfeldt-Jakob disease (CJD) is a rapidly progressive, fatal
neurodegenerative disorder believed to be caused by an abnormal
cellular glycoprotein known as a prion protein.
 Also known as transmissible spongiform encephalopathy (TSE) so
named due to the microscopic appearance of the infected brain
Spongiform changes observed in the gray matter on stained brain
biopsy. This is the most definitive test for diagnosis
 Protein product 14-3-3 can be tested for in CSF
It is a biproduct of cell death in the brain
Not specific but helpful in ruling in/out diagnosis
 Safety important to prevent transmission
• Standard Precautions
• Use disposable equipment when possible
Spongiform
change in the
Gray matter
69

70. 70