1. Viral Infections Terry Kotrla, MS, MT(ASCP)BB

2. Herpes Virus Group Produce a variety of diseases. Produce a variety of diseases. May result in sub-clinical infections May result in sub-clinical infections May be reactivated under appropriate conditions. May be reactivated under appropriate conditions.

3. Herpes Virus Group We will discuss the following: 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. Epstein-Barr virus African or Burkitt’s Lymphoma African or Burkitt’s Lymphoma –malignant B-cell neoplasm –presents as a rapidly growing tumour of the jaw, face or eye –grows very quickly, and without treatment most children die within a few months –Epstein-Barr virus (EBV) has been strongly implicated

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

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

8. B-Cell Lymphoma In most individuals infected with EBV, the virus is present in the B-cells, which are normally controlled by T-lymphocytes 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. 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. EBV induced B cell lymphomas are most prevalent in immunocompromised patients.

9. Oral Hairy Cell Leukoplakia Viral infection of the oral cavity. Viral infection of the oral cavity. Indicator of HIV infection as well as of a person's lessening or weakening immunity Indicator of HIV infection as well as of a person's lessening or weakening immunity

10. Infectious Mononucleosis 4 to 7 week incubation Acute self-limiting infection of the RE system Enlarged lymph nodes in the neck. Sore throat, fever, rash Malaise, lethargy, extreme tiredness Liver and spleen involvement and enlargement Hematology: High WBC, over 20% atypical reactive lymphocytes also known as Downey cells.

11. Infectious Mononucleosis Downey cells may be present Downey cells may be present

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

13. 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.

14. Paul Bunnell Test The original Paul-Bunnell test was a simple titration of sheep cell agglutinins but this procedure was subsequently modified in order to distinguish between sheep cell agglutinins formed in IM and the Forssman-type antibodies found in normal serum, serum sickness and in certain other conditions. The original Paul-Bunnell test was a simple titration of sheep cell agglutinins but this procedure was subsequently modified in order to distinguish between sheep cell agglutinins formed in IM and the Forssman-type antibodies found in normal serum, serum sickness and in certain other conditions. Tissues rich in Forssman antigen (guinea pig kidney) absorb Forssman antibodies but do not affect the heterophil antibodies in IM. Tissues rich in Forssman antigen (guinea pig kidney) absorb Forssman antibodies but do not affect the heterophil antibodies in IM. Heterophil antibodies are absorbed by beef cells, Heterophil antibodies are absorbed by beef cells, Forssman hapten is a glycolipid usually associated with a protein, the determinant being largely carbohydrate and therefore heat stable. Forssman hapten is a glycolipid usually associated with a protein, the determinant being largely carbohydrate and therefore heat stable.

15. Davidsohn Differential The principle behind the Paul-Bunnell-Davidsohn test is that the two types of sheep agglutinins are distinguished by titrating them before and after absorption with guinea pig kidney and ox cells. The principle behind the Paul-Bunnell-Davidsohn test is that the two types of sheep agglutinins are distinguished by titrating them before and after absorption with guinea pig kidney and ox cells. Patients serum containing antibodies due to IM is added to guinea pig kidney cells. These antibodies are not absorbed by the kidney cells. These antibodies then react with Beef (Ox) red blood cells which causes agglutination and is a positive test for IM. Patients serum containing antibodies due to IM is added to guinea pig kidney cells. These antibodies are not absorbed by the kidney cells. These antibodies then react with Beef (Ox) red blood cells which causes agglutination and is a positive test for IM. Patients serum containing Forssman antibodies are added to guinea pig kidney cells. Antibodies are absorbed by the kidney cells. These antibodies are then allowed to react with Beef red blood cells which does not cause agglutination. This is a positive test for Forssman antigens. Patients serum containing Forssman antibodies are added to guinea pig kidney cells. Antibodies are absorbed by the kidney cells. These antibodies are then allowed to react with Beef red blood cells which does not cause agglutination. This is a positive test for Forssman antigens.

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

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

18. 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 Not diagnostic, must look at total clinical picture.

19. 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

20. Cytomegalovirus Transmission occurs from person to person. Close intimate contact Close intimate contact –Sexual contact –Pernatally –Breast milk –Organ transplant –Blood transfusion

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

22. CMV 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.

23. CMV 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

24. Microscopic examination of biopsy specimens

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

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

27. 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

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

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

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

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

32. Neonatal Herpes

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

34. 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

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

37. Chicken Pox

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

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

40. 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

41. Shingles

42. Shingles

43. Shingles

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

45. 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.

46. Rubella

47. Congenital rubella Congenital Rubella Syndrom most serious. Congenital Rubella Syndrom most serious. Fetus infected during first trimester. 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.

48. Rubella Syndrome

49. 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.

50. Laboratory Testing - Rubella Methods include: – –hemagglutination inhibition, – –passive hemagglutination, – –neutralization, – –hemolysis in gel, – –complement fixation, – –fluorescence immunoassay, – –RIA, – –ELISA and – –latex agglutination.

51. 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

52. Rubeola Symptoms include – –irritability, – –runny nose, – –eyes that are red and sensitive to light, – – hacking cough, and – –high fever

53. Rubeola Fever peaks with the appearance of the rash. Rash typically begins on the forehead, then spreads downward over the 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

54. Rubeola

55. Complications Croup bronchitis bronchiolitis pneumonia conjunctivitis myocarditis Hepatitis encephalitis

56. 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 Most severe in adults

57. Measles vaccine Live attenuated Live attenuated DO NOT give to: DO NOT give to: – –pregnant women, – –persons with active tuberculosis, – –leukemia, – –lymphoma, – –depressed immune systems. – –People with egg allergies

58. 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.

59. 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 and – –ELISA.

60. 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.

61. Mumps Single stranded RNA virus. Mumps is transmitted by direct contact with saliva and discharges from the nose and throat iIncubation 16-18 days. glands. Virus can infect many parts of the body, especially the parotid salivary glands.

62. 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

63. Mumps

64. Mumps

65. Mumps - 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

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

67. 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.