1. DNA-VIRUSES. ADENOVIRUSES. HERPESVIRUSES.
Chair of Microbiology, Virology, and Immunology
DNA-VIRUSES. ADENOVIRUSES. HERPESVIRUSES.

2. ADENOVIRUS (ADV)
DNA viruses first isolated from adenoidal tissue in 1953
Family Adenoviridae
Genus Mastadenovirus
There names
adenoid degeneration viruses
adenoid-pharingeal conjunctival viruses
acute respiratory desease agents.

3. DNA-viruses

4. Adenoviruses
Non-enveloped DNA virus
70-90 nm in size
Linear ds DNA genome with core proteins

6. Adenovirus- Properties
Stable in the environment
Relatively resistant to disinfection (Alcohol, chlorhexidine, detergents)
Stable in GI tract- can withstand low pH, bile acids and proteolytic enzymes

7. Dodecon: Hemagglutinin made up of 12 pentons
with their fibers.

8. ICOSAHEDRAL CAPSID
A polyhedron with 20 equilateral faces and 12 vertices
capsomers
ring or knob-shaped units made of 5 or 6 protomers
pentamers (pentons) – 5 subunit capsomers
hexamers (hexons) – 6 subunit capsomers 8

9. Adenoviral proteins
structural
non-structural
According to the time of synthesis in the cell:
early
late
They are differed according to their physical and chemical properties, antigenic specifity. There are capsid and inner proteins. 12 polypeptides (m.w D) are identifyded.

12. ADENOVIRUS - Classification
Subgroups- 6 subgroups (A-F), based on hemagglutination
Serotypes (human)
Common serotypes:- 1-8, 11, 21, 35, 37, 40
Enteric Adenoviruses belong to subgroup F

13. Human Adenoviruses Types
Subgenus
Serotypes A
12, 18, 31 B
3, 7, 11, 14, 16, 21, 34, 35 C
1, 2, 5, 6 D
8, 9, 10, 13, 15, 17, 19, 20, 22-30, 32, 33, 36-39, 42-47 E 4 F
40-41

16. Adenoviruses in the cells

17. Pathogenesis and Replication
Infects mucoepithelial cells of respiratory, GI and GU tracts
Enter via epithelium, replicate and spread to lymphoid tissue
Viremia occurs
Secondary involvement of viscera

18. Pathogenesis and Replication
Fiber protein determines target cell specificity and attachment
Viral DNA enters host cell nucleus
Early and late phases of replication
Error-prone process
Inclusion bodies in nucleus

19. Functional Definition
Adenovirus-Host Cell Interaction
Type of Interaction
Functional Definition
Biologic System
Productive infection
Complete replication of infectious virions
Cultured human cells
Abortive infection
Synthesis of viral gene products without production of infectious virions
Cultured humster or monkey cells
Semipermissive infection
Complete replication with low yields of infectious virions
Cultured rat cells
Malignant transformation
Associated with integration of viral DNA and differential viral and cellular gene expression
Cultured rodent cells
Tumor induction
Newborn hamsters (mice)
Viral latency
Persistant of viral genome
Human tonsils

20. EPIDEMIOLOGY Endemic, epidemic and sporadic infections
Many infections are sub-clinical
Outbreaks:
Military recruits ( 10% basic trainees in 1st week)
swimming pool users, hospitals,
residential institutions, day care settings

21. TRANSMISSION Droplets Fecal-oral route
Direct and through poorly chlorinated water
Fomites

22. Respiratory Eye Genitourinary Gastrointestinal Others
CLINICAL SYNDROMES
Respiratory
Eye
Genitourinary
Gastrointestinal
Others

23. Acute Respiratory Disease
Fever
Tracheobronchitis
Pneumonia
Children and adults
Epidemics in military recruits
Types 4 and 7 most frequently

24. Adenoviral serotypes and associated diseases
Subgroup
Serotype
Clinical syndromes A 31
Infantile gastroenteritis* B
3, 7, 21
Upper respiratory disease, pneumonia, pharyngoconjunctival fever C
1, 2, 5
Hemorrhagic cystitis, interstitial nephritis D
8,19,37
Epidemic keratoconjuctivitis E 4
Upper respiratory disease, pneumonia F
40, 41
Infantile gastroenteritis
Remaining serotypes are infrequently isolated or not clearly associated with disease.
* Association with gastroenteritis not as firmly established as with types 40 and 41.

25. Adenoviral diseases according to patient population
Clinical syndromes
Infants
Pharyngitis, coryza, otitis media
Pneumonia
Diarrhea
Children
Upper respiratory disease
Pharyngoconjunctival fever
Diarrhea, mesenteric adenitis
Hemorrhagic cystitis
Adults
Acute respiratory disease
Epidemic keratoconjunctivitis
Immunocompromised
Gastroenteritis, hepatitis
Hemorrhagic cystitis, interstitial nephritis
Meningoencephalitis

26. Pharyngoconjunctival fever
Headache, fever, malaise
Conjunctivitis and Pharyngitis
Cervical adenopathy, rash and diarrhea also
Main adenovirus types: 3, 4, 7, 14
Epidemics in summer months
Contaminated water in swimming pools,
fomites

27. Adenoviral Infections of the eye
Epidemic Keratoconjunctivitis (EKC)
Acute follicular conjunctivitis
Pharyngoconjunctival fever
Early conjunctivitis (left) and Bilateral conjunctivitis (right)

28. Epidemic Keratoconjunctivitis
Incidence in summer
Types 8, 19, 37
Outbreaks- in situations of close contact (e.g., schools, hospitals, camps, nursing homes, workplaces)
Spread via droplets and contaminated water (ophthalmologic solutions and equipment, swimming pools), fomites, hands

29. EKC-Clinical features
SYMPTOMS
Pink/red eye
Irritation, tearing, foreign-body sensation
Ocular pain
Photophobia
Fever, malaise
Respiratory symptoms
SIGNS
Conjunctival injection, ecchymosis
Corneal injection (limbus)
Diffuse→focal epithelial keratitis
↓Visual acuity (subepithelial corneal opacities)
Ipsilateral pre-auricular lymphadenopathy

30. ADENOVIRAL INFECTIONS - Genitourinary system
Acute hemorrhagic cystitis
fever, dysuria, hematuria
Types 11, 7, 4, 21, 1
More common in boys
Others
Orchitis, nephritis, cervicitis with ulcerated vesicular lesions, urethritis
Types 2, 8, 19, 37

31. Other Infections due to Adenovirus
Myocarditis
Pericarditis
Meningitis
Rash
Arthritis

32. Adenovirus infections in Immunocompromised hosts
Disseminated, severe and often fatal infections
Due to new infection or reactivation of latent virus
Prolonged infections with prolonged viremia and viral shedding
Necrotizing pneumonia, hepatitis, rash, DIC, CNS involvement

33. DIAGNOSIS OF ADENOVIRAL INFECTIONS
Variety of clinical specimens depending on clinical syndrome-NP, conjunctival, stool, urine, tissue, etc.
Transport in viral transport media
Isolation from pharyngeal/nasal site within 3 days of symptom onset ( 3 weeks for EKC)

34. Methods for ADV diagnosis
Culture in HeLa, HEK cell lines
Shell vial cell culture
DFA/EIA insensitive except types 40, 41
PCR, nucleic acid probes esp. myocarditis
EM and Immune EM
Histology characteristic intranuclear inclusions
Serology serum titer corresponding to culture isolate
Diagnosis-Enteric adenoviruses
Isolation requires special media-Graham 293
ELISA for rapid detection is available

35. Therapy for ADV Infections
Antiviral Cidofovir – limited data
Immunologic Pooled immunoglobulin esp.
pediatric – associated serotypes
Activated T cell infusion –
anecdotal

36. ADENOVIRUS VACCINE Oral live attenuated vaccine Strains 4, 7
Used in military recruits
Manufacture of vaccine was halted in 1996
Lapse in immunization was associated with outbreaks in military recruits

37. Herpesvirus Family There are about 80 herpesviruses.
Herpes simplex virus 1,
Herpes simplex virus 2,
Cytomegalovirus,
Herpes zoster virus-Varicella virus),
Epstein-Barr virus,
Herpesvirus 6,
Herpesvirus 7. 8
Viruses can be isolated from cows, pigs, hens

38. All herpesviruses have a core of double-stranded DNA surrounded by a protein coat that exhibits icosahedral symmetry and has 162 capsomeres. The nucleocapsid is surrounded by an envelope. The enveloped form measures nm; the "naked" virion, 100 nm. The double-stranded DNA (MW xlO6) has a wide range of guanine + cytosine content in different herpesviruses, There is little DNA homology among different herpesviruses, except her­pes simplex types 1 and 2.

39. Herpesviruses

40.  (Betaherpesvirinae),
Classification
Three subfamily:
 (Alphaherpesvirinae),
 (Betaherpesvirinae),
 (Gammaherpesvirinae)
Alphaherpesvirinae: a lot of hosts, short reproductive cycle, effective cells (infected ones) destroying, latent form of existence in ganglia.
Simplexvirus (HSV-1, HSV-2) і Poikilovirus.

41. Betaherpesvirinae:
Limited spectrum of hosts, reproductive cycle is long.
Infected cells increase their sizes, formation of persistence infections in cell cultures. Virus exist in latent form in limphoreticular cells, secretory glands, kidneys and so on.
Genera:
Cytomegalovirus
Muromegalovirus (mice cytomegalovirus)

42. Gammaherpesvirinae (EBV, HV-6, 7, 8 MDV):
In vitro viruses replicate in liphoblast cells, cause lytic infection in epithelioid cells fibroblasts.
Viruses have tropism both Т-, and В-lymphocytes, but as a rool infection process is stopped at prelytic stage, so productive cases are absent.

43. HERPES SIMPLEX (Human Herpesvirus 1 and 2)
(Herpes labialis, Herpes genitalis, and many other syndromes).
Infection with herpes simplex virus (herpesvirus hominis) may take several clinical forms. The infection is most often inapparent. The usual clinical man­ifestation is a vesicular eruption of the skin or mucous membranes. Infection is sometimes seen as severe keratitis, meningoencephalitis, and a disseminated illness of the newborn.
Properties of the Virus. The envelope is derived from the nuclear membrane of the infected cell. It contains lipids, carbohydrate, and protein and is removed by ether treatment. The double-stranded DNA genome is linear (MW x 106).

44. Types 1 and 2 show 50% sequence homology
Types 1 and 2 show 50% sequence homology. Treatment with restriction endonucleases yields characteristically different cleavage patterns for type 1 and 2 viruses and even for different strains of each type. This "fingerprinting" of strains allows epidemiologic tracing of a given strain, whereas in the past, the ubiquitousness of herpes simplex virus made such investigations impossible.

45. Herpesvirus. Several proteins have been identified in the virion, The protein I is associated with the viral capsid; the glycoproteins present in bands 0, 2 -3, 8, and 9 (glycoproteins) are associated with the envelope; and a DNA-binding protein (band number 13) is associated with the internal core.

46. Chorionallantoic membrane (right) infected with Herpesvirus

47. Viral cytopathogenic effect (right)
The plaques produced by herpesvirus type 2 are larger than the tiny plaques produced by type 1 virus. The virus grows readily and produces plaques in almost any cell culture. Infected cells develop inclusion bodies and then undergo necrosis (cytopathic effect).
In Chinese hamster cells, which contain 22 chromosomes, the virus causes breaks in region 7 of chromosome No. 1 and in region 3 of the X chromosome. The Y chromosome is unaffected.
Viral cytopathogenic effect (right)

49. Differentiation of Types 1 and 2
Herpes simplex virus types 1 and 2 cross-react serologically but may be distinguished by a number of tests: (1) The use of type-specific antiserum prepared by adsorption of the viral antiserum with heterotypically infected cells or by inoculation of rabbits with individual type-specific proteins. (2) The greater temperature sensitivity of type 2 infectivity. (3) Preferential growth in different cell species. (4) Restriction enzyme patterns of virus DNA molecules. (5) Differences in the polypeptides produced by type 1 and type 2.
Oncogenic Properties: After inactivation of their lytic capabilities by ultraviolet irradiation or other means, herpesvirus types 1 and 2 can cause transformation of cultured hamster cells, which may induce tumours when inoculated into newborn hamsters. Viral genetic information can be demonstrated in the tumour cells.

50. Herpes Simplex Virus
HSV is spread by contact, as the virus is shed in saliva, tears, genital and other secretions.
Primary infection is usually trivial or subclinical in most individuals. It is a disease mainly of very young children ie. those below 5 years.
About 10% of the population acquires HSV infection through the genital route and the risk is concentrated in young adulthood.
Following primary infection, 45% of orally infected individuals and 60% of patients with genital herpes will experience recurrences.
The actual frequency of recurrences varies widely between individuals. The mean number of episodes per year is about 1.6.

51. Pathogenesis
During the primary infection, HSV spreads locally and a short-lived viraemia occurs, whereby the virus is disseminated in the body. Spread to the to craniospinal ganglia occurs.
The virus then establishes latency in the craniospinal ganglia.
The exact mechanism of latency is not known, it may be true latency where there is no viral replication or viral persistence where there is a low level of viral replication.
Reactivation - It is well known that many triggers can provoke a recurrence. These include physical or psychological stress, infection; especially pneumococcal and meningococcal, fever, irradiation; including sunlight, and menstruation.

52. Pathogenesis of herpetic infection

53. Herpesvirus Type 1
The clinical entities attributable to herpesvirus type 1 include the following:
1. Acute herpetic gingivostomatitis (aphthous stomatitis, Vincent's stomatitis). This is the most common clinical entity caused by primary infections with type 1 herpesvirus. It occurs most frequently in small children (1-3 years of age) and includes extensive vesiculoulcerative lesions of the mucous membranes of the mouth, fever, irritability, and local lymphadenopathy. The incubation period is short (ab­out 3-5 days), and the lesions heal in 2-3 weeks.
2. Eczema herpeticum (Kaposi's varicelliform eruption). This is a primary infection, usually with herpesvirus type 1, in a person with chronic eczema. In this illness, there may be extensive vesiculation of the skin over much of the body and high fever. In rare instances, the illness may be fatal.

55. 3. Keratoconjunctivitis
3. Keratoconjunctivitis. The initial infection with herpesvirus may be in the eye, producing severe keratoconjunctivitis. Recurrent lesions of the eye appear as dendritic keratitis or corneal ulcers or as vesicles on the eyelids. With recurrent keratitis, there may be progressive involvement of the corneal stroma, with permanent opacification and blindness.
4. Encephalitis. A severe form of encephalitis may be produced by herpesvirus. In adults, the neurologic manifestations suggest a lesion in the temporal lobe. Pleocytosis (chiefly of lymphocytes) is present in the cerebrospinal fluid; however, definite diagnosis during the illness can usually be made only by isolation of the virus (or by demonstrating viral antigens by immunofluorescence) from brain tissue obtained by biopsy or at post-mortem. The disease carries a high mortality rate, and those who survive often have residual neurologic defects.

58. 5. Herpes labialis (cold sores, herpes febrilis)
5. Herpes labialis (cold sores, herpes febrilis). This is the most common recurrent disease produced by type 1. Clusters of localized vesicles occur, usually at the mucocutaneous junction of the lips. The vesicle ruptures, leaving a painful ulcer that heals without scarring. The lesions may recur, repeatedly and at various intervals of time, in the same location. The permanent site of latent herpes simplex virus is the trigeminal ganglion.

59. Congenital herpes

60. Herpesvirus Type 2
1. Genital herpes (herpes progenitalis): vesiculoulcerative lesions of the penis of the male or the cervix, vulva, vagina, and perineum of the female. The lesions may be associated with fever, malaise, and inguinal lymphadenopathy. In women with herpesvirus antibodies, only the cervix or vagina may be involved, and the disease may therefore be asymptomatic.
Type 2 virus remains latent in lumbar and sacral ganglia. Changing patterns of sexual behaviour are reflected by an increasing number of type 1 virus isolations from genital lesions and of type 2 from facial lesions, presumably as a result of oral-genital sexual activity.

61. Genital herpes

62. HV, immune fluorescence test

63. VARICELLA-ZOSTER VIRUS (Human Herpesvirus 3)
VARICELLA (Chickenpox) ZOSTER (Herpes Zoster, Shingles, Zona)
Varicella (chickenpox) is a mild, highly infectious disease, chiefly of children, characterized clinically by a vesicular eruption of the skin and mucous membranes. However, in immunocompromised children the disease may be severe. The causative agent is indistinguishable from the virus of zoster.
Zoster (shingles) is a sporadic, incapacitating disease of adults (rare in children) that is characterized by an inflammatory reaction of the posterior nerve roots and ganglia, accompanied by crops of vesicles (like those of varicella) over the skin supplied by the affected sensory nerves.

64. VARICELLA-ZOSTER VIRUS

65. Varicella, pathogenesis

66. Varicella

67. Herpes zoster pathogenesis

68. Herpes zoster

69. VARICELLA-ZOSTER
VIRUS, IFT

70. CYTOMECALOVIRUS (Human Herpesvirus 5) (Cytomegalic Inclusion Disease)
Cytomegalic inclusion disease is a generalized infection of infants caused by intrauterine or early postnatal infection with the cytomegaloviruses. The disease causes severe congenital anomalies in about 10,000 infants in the USA per year. Cytomegalovirus can be found in the cervix of up to 10% of healthy women.

71. Cytomegalovirus in the cells

72. Retina infected by Cytomegalofirus
Cytomegalovirus, IFT

73. EB HERPESVIRUS (Human Herpesvirus 4)
(Infectious Mononucleosis, Burkitt's Lymphoma,
Nasopharyngeal Carcinoma)
EB (Epstein-Barr) virus is the causative agent of infectious mononucleosis and has been associated with Burkitt's lymphoma and nasopharyngeal carcinoma. The virus is an antigenically distinct herpesvirus.
Properties of the Virus. Morphology: EB virus is indistinguishable in size and structure from other herpesviruses.
Antigenic Properties: EB virus is distinct from all other human herpesviruses. Many different EB virus antigens can be detected by CF, immunodiffusion, or immunofluorescence tests. A lymphocyte-detected membrane antigen (LYDMA) is the earliest-detected virus-determined antigen. EBNA is a complement-fixing nuclear antigen. Early antigen (EA) is formed in the presence of DNA inhibitors and membrane antigen (MA), the neutralizing antigen, is a cell surface antigen. The virus capsid antigen (VCA) is a late antigen representing virions and structural antigen.

74. EBV

75. Infectious mononucleosis
Angina in patient with infectious mononucleosis

77. Burkitt's Lymphoma

78. Nasopharyngeal Carcinoma -

79. Human Herpesvirus 6 and 7
Pathogenesis. The reservoir and mode of transmission of human herpesvirus 6 and 7 are not well understood at the present time. It should be noted that high prevalence of antibodies early in life would implicate transmission within the home from oropharyngeal secretions; however, this has not yet been documented.
Epidemiology. The epidemiology of human herpesvirus 6 and 7 is poorly understood at present. Loss of transplacental antibodies, followed by acquisition of antibodies early in life, implies horizontal transmission within the home environment. By the age of 5, antibodies are present in virtually 100 per cent of the population to both of these viruses. Human herpesvirus 6 exists as type A and B. The type A variant was the original isolate being retrieved from an immunocompromised host. The type B variant is associated with roseola. Some investigators suggest that the genetic differences in these two types warrant distinct names; thus, it is conceivable that the International Herpesvirus Nomenclature Committee may designate these agents as distinct.

80. Human Herpesvirus 6

81. Clinical Manifestations
Clinical Manifestations. Human herpesvirus 6, 7, 8 have recently been isolated. Human herpes virus 6 (as has human herpes virus 7, but to a lesser extent), has been associated with exanthem subitum, or roseola. This illness is characterized by days of fever, followed by the appearance of a maculopapular "slapped cheek" rash. In addition, there has been an association between human herpesvirus 6 and rejection of transplanted kidneys, fulminant hepatitis and infections of the central nervous system.
Kaposi's Sarcoma Herpesvirus (HHV-8). Recently, a new herpesvirus has been associated with Kaposi's sarcoma and AIDS-related lymphomas of organ cavities. The DNA of this virus is partially homologous to the DNA of Epstein-Barr virus and that of herpesvirus saimiri. This virus immortalizes B lymphocytes. Isolation of the virus has yet to be achieved.

82. THE END