1. Unit 4 Part 5 Viral Infections
Terry Kotrla, MS, MT(ASCP)BB

2. Herpes Virus Group Include 8 viruses that cause disease.
May result in sub-clinical infections
Capable of establishing latent infection.
May be reactivated under appropriate conditions.

3. Herpes Virus Group We will discuss the following: Epstein-Barr virus
Cytomegalovirus
Herpes simplex virus type I and II
Varicella-zoster virus

4. Epstein-Barr Virus (EBV)
Spread through oral transmission
Cause of Infectious Mononucleosis.
Other Diseases include:
African or Burkitt’s lymphoma
Nasopharyngeal carcinoma
B cell lymphoma

5. Infectious Mononucleosis
Acute self-limiting infection of the RE system
Four to 7 week incubation
Enlarged lymph nodes in the neck.
Sore throat, fever, rash
Malaise and extreme tiredness
Liver and spleen involvement and enlargement.

6. Infectious Mononucleosis - Lab
Hematology:
High WBC,
Over 20% atypical reactive lymphocytes also known as Downey cells.
Positive mono (heterophile antibody) test.
Negative mono test do tests for antibodies to EBV associated antigens.

7. Infectious Mononucleosis
Downey cells may be present

8. Heterophile Antigens/Antibodies
Heterophile antigens are a group of similar antigens found in unrelated animals, i.e., man, sheep, horse, dog cat, mouse.
Heterophile antibodies produced against heterophile antigens of one species will cross react with others.

9. Heterophile Antigens/Antibodies
Forssman antigen is an example of a heterophile antigen and is found on the RBCs of many species (guinea pig, dog, cat, mouse, sheep, fowl, horse)
Forssman antibodies formed against Forssman antigens will agglutinate sheep RBCs.

10. Paul Bunnell Test Simple titration of sheep cell agglutinins.
Serial dilutions of patient serum.
Add sheep RBCs.
Positive indicates presence of heterophile antibodies.
Not specific for IM, screening test only.

11. Davidsohn Differential
Used to determine heterophile antibody present.
Add patient serum to one tube with guinea pig kidney and another with beef red blood cells.
Beef RBCs have IM antigen.
Guinea pg kidney have Forssman antigen.
Harvest serum and test with sheep cells.

12. Davidsohn Differential
Davidsohn Differential * To be considered absorbed there must be greater than a three tube difference between the presumptive titer and the differential titer.

13. Davidsohn Differential
Advantages
When properly performed, this test is specific for Infectious Mononucleosis
False-positive results are rare.
Disadvantages
Davidsohn Differential test is very time consuming and burdensome.

14. Infectious Mono Slide Tests
It was discovered that horse RBCs possess antigens which react with the antibody associated with IM.
Patient serum mixed with horse RBCs, agglutination is positive.
Latex agglutination, coat particles with EBV antigens.
Not diagnostic, must look at total clinical picture.

15. EBV Antigens Early antigen (EA) Capsid antigen (VCA)
Membrane antigen (MA)
Nuclear antigen (EBNA)

16. EBV Specific Antibodies
EBV specific antibodies may be measured.
Pattern of appearance of EBV antigens.
Most valuable is IgM antibody to viral capsid antigen (VCA), indicates a current infection (best marker), lasts about 12 weeks.
Can also detect anti-early antigen (EA) (recent infection) and anti EB nuclear antigen (EBNA) (older infection).
ELISA and IFA most commonly used

17. EBV Antibody Response Stage of Disease Anti-VCA Heterophile Antibody
IgM
IgG
Anti-EBNA
Acute
Pos or Neg
Pos
Neg
Past infection

18. African or Burkitt’s Lymphoma
EBV has been strongly implicated
Malignant B-cell neoplasm
presents as rapidly growing tumor of the jaw, face or eye
grows very quickly, and without treatment most children die within a few months
Rare in US, equatorial Africa.
Also associated with HIV infection.

19. African or Burkitt’s Lymphoma
Although BL is a very rapidly growing tumour it responds well to treatment.
Three pictures: before treatment, 3 days and 6 days after treatment

20. Nasopharyngeal Carcinoma
Endemic in South China, Africa, Arctic Eskimos
This is a malignant tumour of the squamous epithelium of the nasopharynx.
100% contain EBV DNA
Rates are less than 1 per 100,000 in most populations
Nasopharyngeal carcinomas are found in association with reactivation of latent Epstein-Barr Virus. 
The exact mechanisms of association are unknown

21. B-Cell Lymphoma
In most individuals infected with EBV, the virus is present in the B-cells, which are normally controlled by T-lymphocytes
When T-cell deficiency exists, one clone of EBV-infected B-lymphocytes escapes immune surveillance to become autonomously proliferating.
EBV induced B cell lymphomas are most prevalent in immunocompromised patients.

22. Oral Hairy Cell Leukoplakia
HIV infection stimulates reactivation of pre-existing, latent EBV infection.
Causes viral infection of the oral cavity.
Indicator of HIV infection as well as of a person's lessening or weakening immunity

23. Cytomegalovirus Human Herpesvirus 5 (HHV-5)
Transmission occurs from person to person.
Close intimate contact
Sexual contact
Perinatally
Breast milk
Organ transplant
Blood transfusion

24. CMV Clinical course Symptoms resemble IM
In babies may cause life threatening illness
Patients with deficient immune systems
AIDS patients
Transplant patients

25. Cytomegalovirus Perinatal transmission occurs in 10%.
If infected may cause multitude of symptoms
Petechiae
Jaundice
Hepatosplenomegaly
Neurological abnormalities
Moratlity rate 5 percent
Survivors may exhibit hearing loss, visual impairment and mental retardation.

26. CMV Immunologic response
Test for CMV antibody using paired serum samples
IgM antibodies produced against early and intermediate-early (IE) CMV antigens, last for 3 to 4 months.
IgG appear shortly after and peak at 2 to 3 months.

27. CMV Laboratory Diagnosis
Range from culture and cytologic techniques to DNA probes, PCR and serologic techniques.
Detection of antibodies indicator of recent or active infection.
Viral cultures

28. Microscopic examination of biopsy specimens

29. CMV Lab Diagnosis Detection of CMV antigen in cells using IFA
ELISA to detect antibody to CMV
Other
fluorescence assays,
indirect hemagglutination, and
latex agglutination
False positives can occur due to RA and Epstein-Barr antibodies

30. Herpes Simplex Virus (HSV)
Most exposed in childhood
Possesses viral latency – hibernation
Two types: HSV-1 and HSV-2

31. HSV-1 Transmitted from person to person by saliva or direct contact.
Cold sores around the mouth most common.
Reactivation - may have several episodes of cold sores during a lifetime

32. HSV-1 Symptoms Blister forms, breaks, crusts over
tingling
Numbness
Itching
Blister forms, breaks, crusts over
Reactivation usually caused by stress.
Conjunctivitis, keratitis and herpetic whitlow may occur.

33. HSV-2 Results in Herpes genitalis - lesions BELOW the waist.
Transmitted intimate sexual contact or perinatally.
Symptoms
Pain
Tenderness
Itch
Fever
Headache
Lymphadenopathy
Malaise

34. HSV-2 Blisters appear Painful, lasts 1-3 weeks
Males – penis
Females – vagina and cervix
Both – thighs buttocks
Painful, lasts 1-3 weeks
Virus lies dormant in nearby nerves and reactivated.

35. HSV – 2 Can be fatal in infants
Woman with active infection needs C-section.
Infants with localized infections have 70% mortality rate
Disseminated neonatal herpes most lethal form.

36. Neonatal Herpes

37. Laboratory Testing for HSV
Recovery of virus from culture
Direct examination of cells from lesion using IF or immunoperoxidase stain
DNA probes
ELISA
Latex agglutination
RIA
Indirect IF
Serology NOT very useful

38. Varicella-Zoster Virus
Two different manifestations of the same virus.
Varicella is the primary infection, causes chicken pox
Herpes Zoster causes shingles and is due to reactivation of the latent virus

39. Varicella Fever and vesicular exanthema
Small, itchy blisters surrounded by inflamed skin.
Begins as one or two lesions and spreads.
Number of lesions vary greatly.
Blister dries out and forms a scab.

41. Chicken Pox

42. Chicken Pox Secondary complications due to infection most common.
May also result in pneumonia, encephalitis and hepatitis.
Very serious for immunocompromised children
Vaccine now available

43. Shingles Chicken pox – virus goes latent Reactivated later in life
Weakened immune system
Aging
Other factors

44. Shingles
The typical rash of shingles begins as redness(erythema) followed by the appearance of blisters.
Eruptions follow the path of an infected nerve.
The trunk is the area affected in 50% to 60% of cases.
Skin may be extremely sensitive to touch

45. Shingles

46. Shingles

47. Shingles

48. VZV Laboratory Testing
Important to distinguish VZV from other infections
PCR
Direct Fluorescent Antibody staining
Viral culture
IgG and IgM antibody test by ELISA

49. Rubella Virus
RNA virus with 3 major structural proteins, E1, E2, and C.
Incubation 2- 3 weeks
Highly contagious, spread through respiratory tract.
Causes German measles
Rubella vaccine has resulted in 99% decline in infections.

50. Rubella Symptoms Mild and difficult to notice Mild fever Headache
Stuffy or runny nose
Red eyes
Enlarged and tender lymph nodes
Pink rash begins on face, spreads to trunk then arms and legs, disappears in same order.
Aching joints

51. Rubella

52. Congenital Rubella Congenital Rubella Syndrome most serious.
Fetus infected during first trimester.
Result in miscarriage or stillbirth,
Live-born serious birth defects or dying.
20% of the children born after such an infection suffer the severe congenital abnormalities
10-20% of these children die within the first year of life.
Rubella vaccine contraindicated during pregnancy.

53. Rubella Syndrome

54. Rubella Lab testing IgG and IgM antibodies may form at same time
IgM antibodies persist for 4 to 5 weeks, IgG for life.
Performed primarily for diagnosis of acquired infections and to determine immune status of pregnant patients.
Some tests detect IgG antibodies, other IgM.

55. Rubella Laboratory Testing
Methods include:
hemagglutination inhibition
passive hemagglutination
neutralization
hemolysis in gel
complement fixation
fluorescence immunoassay
RIA
ELISA
latex agglutination.

56. Rubeola Single stranded RNA virus best known for its typical skin rash
Primarily respiratory infection
Incubation approximately 10 days, ranges from 8-13.
Rash appears at about day 14.
Airborne precautions

57. Rubeola Symptoms include Irritability Runny nose
Eyes that are red and sensitive to light,
Hacking cough, and
High fever
Swollen lymph nodes

58. Rubeola Fever peaks with the appearance of the rash.
Red spots with tiny grayish white heads appear on inside of cheeks at back of mouth.
After 18 hours spots disappear and rash develops, typically begins on forehead, then spreads downward over face, neck, and body.
Rash appears on face first and consists of large flat red to brown blotches that often flow into one another.
Rash fades in the same order that it appeared

59. Rubeola

60. Rubeola Complications
Croup
Bronchitis
Bronchiolitis
Pneumonia
Conjunctivitis
Myocarditis
Hepatitis
Encephalitis

61. Rubeola More susceptible to ear infections or pneumonias.
Disease can be severe, with bronchopneumonia or brain inflammation
May lead to death in approximately 2 of every 1,000 cases.
Most severe in adults.

62. Measles vaccine
Live attenuated-reduce virulence of organism but it is till alive.
DO NOT give to:
pregnant women,
persons with active tuberculosis,
leukemia,
lymphoma,
depressed immune systems.
People with egg allergies

63. Measles vaccine
Occasionally causes side effects in persons with no underlying health problems,
In about 10% of cases there is a fever between 5 and 12 days after vaccination,
In about 5% of cases there is a rash.

64. Rubeola Laboratory Testing
Serology testing provides best means of confirming a measles diagnosis
Methods to detect rubeola antibodies include:
hemagglutination inhibition
endpoint neutralization
complement fixation
IFA
ELISA

65. Laboratory Testing
Diagnosis confirmed by presence of Rubeola specific IgM antibodies antibodies or four-fold rise in IgG antibody titer in paired samples taken after rash to 10 to 30 days later.
IgM test highly depended on time of sample collection with 3-11 days after rash being optimal.
IgM false positive due to RA.

66. Mumps Single stranded RNA virus.
Mumps is transmitted by direct contact with saliva and discharges from the nose and throat
iIncubation days.
Virus can infect many parts of the body, especially the parotid salivary glands.

67. Mumps
Glands usually become increasingly swollen and painful over a period of 1 to 3 days
Pain gets worse
Both the left and right parotid glands may be affected

68. Mumps

69. Mumps

70. Mumps - Complications Inflammation and swelling of the brain
Mumps in adolescent and adult males may also result in the development of orchitis
May affect the pancreas or, in females, the ovaries
Infection in pregnant women may result in increased risk for fetal death

71. Laboratory Testing complement fixation hemagglutination inhibition
hemolysis-in-gel
neutralization assys
IFA and
ELISA

72. Laboratory Testing
Current or recent infections indicated by presence of specific IgM antibody in single sample which can be detected within 5 days of illness.
Fourfold rise in specific IgG antibody in 2 samples collected during acute and convalescent phases
Fluorescent antibody staining for mumps antigens
Cross-reactivity between antibodies to mumps and parainfluenza viruses has been reported in tests for IgG, not a problem since symptoms differ.

73. The End