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  1. 1. cytomegalovirus Cytomegalovirus
  2. 2. • Cytomegaloviruses are herpesviruses • The name for the classic cytomegalic inclusion disease derives from the property for massive enlargement of cytomegalovirus-infected cells. • Cytomegalic inclusion disease is a generalized infection of infants caused by intrauterine or early postnatal infection with the cytomegaloviruses. • Cytomegalovirus poses an important public health problem because of its high frequency of congenital infections, which may lead to severe congenital anomalies. • Inapparent infection is common during childhood and adolescence. • Severe cytomegalovirus infections are frequently found in adults who are immunosuppressed.
  3. 3. Properties of the Virus • Cytomegalovirus has the largest genetic content of the human herpesviruses. Its DNA genome 240kbp • Cytomegaloviruses are very species-specific and cell type-specific • Human cytomegalovirus replicates in vitro only in human fibroblasts. • Cytomegalovirus replicates very slowly in cultured cells • infection is spread primarily cell-to-cell. • It may take several weeks for an entire monolayer of cultured cells to become involved • Cytomegalovirus produces a characteristic cytopathic effect . Perinuclear cytoplasmic inclusions ,intranuclear inclusions. Multinucleated cells are seen. Many affected cells become greatly enlarged. • Inclusion-bearing cytomegalic cells can be found in samples from infected individuals
  4. 4. Pathogenesis & Pathology I- Normal host • transmitted person-to-person in several different ways, all requiring close contact with virus-bearing material • There is a 4- to 8-week incubation period in normal older children and adults after viral exposure. • The virus causes a systemic infection; it has been isolated from lung, liver, esophagus, colon, kidneys, monocytes, and T and B lymphocytes. • The disease is an infectious mononucleosis-like syndrome, although most cytomegalovirus infections are subclinical. • cytomegalovirus establishes lifelong latent infections. • Virus can be shed intermittently from the pharynx and in the urine for months to years after primary infection . • Salivary gland involvement is common and is probably chronic.
  5. 5. II-mmunosuppressed Hosts • • • • • • Primary cytomegalovirus infections in immunosuppressed hosts are much more severe than in normal hosts. Individuals at greatest risk for cytomegalovirus disease are those receiving organ transplants, those with malignant tumors who are receiving chemotherapy, and those with AIDS. Viral excretion is increased and prolonged, and the infection is more liable to become disseminated. Pneumonia is the most common complication. The host immune response presumably maintains cytomegalovirus in a latent state in seropositive individuals. Reactivated infections are associated with disease much more often in immunocompromised patients than in normal hosts. Although usually less severe, reactivated infections may be as virulent as primary infections.
  6. 6. III-Congenital and Perinatal Infections • Fetal and newborn infections with cytomegalovirus may be severe. • About 1% of live births annually in the United States have congenital cytomegalovirus infections and about 5–10% of those will suffer cytomegalic inclusion disease. • Babies with this disease will exhibit developmental defects and mental retardation.
  7. 7. • The virus can be transmitted in utero with both primary and reactivated maternal infections. • About one-third of pregnant women with primary infection transmit the virus. • Generalized cytomegalic inclusion disease results most often from primary maternal infections. • There is no evidence that gestational age at the time of maternal infection affects expression of disease in the fetus. • Intrauterine transmission occurs in about 1% of seropositive women. Fetal damage seldom results from these reactivated maternal infections; the infection of the infant remains subclinical though chronic . • Cytomegalovirus can also be acquired by the infant from exposure to virus in the mother's genital tract during delivery and from maternal breast milk. In these cases, the infants usually have received some maternal antibody, and the perinatally acquired cytomegalovirus infections tend to be subclinical. • Transfusion-acquired cytomegalovirus infections in newborns will vary, depending on the amount of virus received and the serologic status of the blood donor. • Whether cytomegalovirus is acquired in utero or perinatally, a more chronic infection results—with respect to viral excretion—than when the virus is acquired later in life
  8. 8. Clinical Findings I-Normal Hosts • Primary cytomegalovirus infection of older children and adults is usually asymptomatic but occasionally causes a spontaneous infectious mononucleosis syndrome. The disease is characterized by malaise, myalgia, protracted fever, liver function abnormalities, and lymphocytosis. Cytomegalovirus is estimated to cause 20–50% of heterophil-negative (non-Epstein-Barr virus) mononucleosis cases. • Cytomegalovirus mononucleosis is a mild disease, and complications are rare. Subclinical hepatitis is common. In younger children (under 7 years old), hepatosplenomegaly is frequently observed. • An association has been observed between the presence of cytomegalovirus and restenosis following coronary angioplasty. It is speculated that the virus may be contributing to the proliferation of smooth muscle cells, leading to restenosis.
  9. 9. II-Immunocompromised Hosts • Both morbidity and mortality rates are increased with primary and recurrent cytomegalovirus infections in immunocompromised individuals. • Pneumonia is a frequent complication. Interstitial pneumonitis caused by cytomegalovirus occurs in 10–20% of bone marrow transplant recipients. • Virus-associated leukopenia is common in solid organ transplant recipients; also seen are obliterative bronchiolitis in lung transplants, graft atherosclerosis after heart transplantation, and cytomegalovirus-related rejection of renal allografts. • Cytomegalovirus often causes disseminated disease in untreated AIDS patients; gastroenteritis and chorioretinitis are common problems, the latter often leading to progressive blindness
  10. 10. III-Congenital and Perinatal Infections • Congenital infection may result in death of the fetus in utero . • Cytomegalic inclusion disease of newborns is characterized by involvement of the central nervous system and the reticuloendothelial system. Clinical features include intrauterine growth retardation, jaundice, hepatosplenomegaly, thrombocytopenia, microcephaly, and retinitis. • Mortality rates are about 20%. The majority of survivors will develop significant central nervous system defects within 2 years; severe hearing loss, ocular abnormalities, and mental retardation are common. • About 10% of infants with subclinical congenital cytomegalovirus infection will develop deafness.
  11. 11. • Many women infected previously with cytomegalovirus show reactivation and begin to excrete the virus from the cervix during pregnancy. • At the time of delivery through the infected birth canal, infants may become infected, though they possess high titers of maternal antibody acquired transplacentally. • These infants begin to shed virus at about 8–12 weeks of age. • They continue to excrete the virus for several years but remain healthy. • Acquired infection with cytomegalovirus is common and usually inapparent. The virus is shed in the saliva and urine of infected individuals for weeks or months. Cytomegalovirus may be a cause of isolated pneumonia in infants less than 6 months of age
  12. 12. Immunity • Antibodies to cytomegalovirus occur in most human sera. • Cytomegalovirus-specific antibodies of the IgM, IgA, and IgG classes have all been detected. • Reactivation of latent infection occurs in the presence of humoral immunity. • The presence of antibody in breast milk does not prevent transmission of infection to breastfeeding infants. • Maternal antibody protects more against development of serious disease in the infant than viral transmission.
  13. 13. • 1. Humoral Response : CMV specific IgM antibodies are produced during the primary infection and persists for 3 or 4 months, but are not produced in recurrent infections in immunocompetent individuals. However, immunocompromised individuals may fail to produce IgM with primary infection and 1/3rd of them have IgM detectable with recurrent infection. CMV IgG antibodies are produced at the time of primary infection and persists lifelong. With intrauterine infections, both IgM and IgG are produced by the fetus but the fetal IgG response can only be detected as the passively acquired IgG from the mother is catabolized. Although there is evidence to suggest that the humoral response may be beneficial, eg. immunocompromised patients who fail to develop IgM run a high risk of developing disseminated infection, the exact role of the humoral response remain uncertain. • 2. CMI: CMI is thought to play a key role in the suppression of CMV infection. The test most widely used to measure CMI is the lymphocyte transformation response (LTR) which measures the recognition, not the effector function of T lymphocytes. Most seropositive adults have a positive LTR, whereas few congenitally or perinatally infected infants can respond. This failure of LTR recovers with time and there is a direct correlation between cessation of viruria and acquisition of LTR responsiveness at 3 -5 years of age.
  14. 14. Laboratory Diagnosis Polymerase Chain Reaction and Antigen Detection Assays • PCR assays have replaced virus isolation for routine detection of cytomegalovirus infections. Cell culture methods of viral isolation are too slow to be useful in guiding therapy, particularly in immunosuppressed patients. The PCR assays are designed to detect replicating virus, not latent viral genomes. Blood and urine are most commonly tested. PCR assays can provide viral load data, which appears to be important in predicting cytomegalovirus disease. Monoclonal antibodies against viral antigens can be used to detect virus-positive leukocytes from patients. Isolation of Virus • Human fibroblasts are used for virus isolation attempts. The virus can be recovered most readily from throat washings and urine. In cultures, 2–3 weeks are usually needed for the appearance of cytologic changes, consisting of small foci of swollen, translucent cells with large intranuclear inclusions. The virus stays cell-associated. Serology • Many types of assays can detect cytomegalovirus IgG antibodies, indicative of past infection (and the potential to undergo reactivation). Detection of viral IgM antibodies suggests a current infection. Serologic assays are not informative for immunocompromised patients. Furthermore, serologic techniques cannot distinguish strain differences among clinical isolates.
  15. 15. • 1. Virus Isolation ;- Urine, saliva, blood and biopsy samples can be used for virus isolation. • Urine should be collected a sterile container without additives. • Saliva samples should first be soaked on to a swab which is then broken off into transport medium. • Blood should be collected into a heparinized bottle containing 500 units of heparin. • Tissue biopsies should be placed in sterile plastic containers. The specimens can be treated in the following ways ;• .
  16. 16. • (a) Cell culture - Human embryo lung fibroblasts are most commonly used. The specimen is inoculated into HEL cells and kept for 28 days with a blind passage at 14 days. CMV produces a typical focal cytopathic effect. • (b) DEAFF ( Detection of early antigen fluorescent foci ) ;- This is a method used for the early diagnosis of CMV infection. In immunocompromised patients, a sensitivity of 78% and a specificity of 100%. The specimen is inoculated into cell culture which is examined 24 hours later by immunofluorescence for expressed CMV encoded early proteins. The monoclonal antibodies must be able to cover most, if not all strains of CMV. Rapid culture methods other then the DEAFF tests are also available. • (c) Histopathology - Cytomegalic inclusions can be recognized from biopsy material by the typical "owl 's eyes appearance " • (d) Tissue immunofluorescence - Infected lung and liver cells may be stained by specific anti-CMV antibodies. Broncheolavage specimens can also be examined in this manner. Results of high sensitivity and specificity.
  17. 17. • e) Electron microscopy - Virions in the urine of congenitally infected infants may be visualized by EM in up to 80% of cases. However this is of no real value as rapid diagnosis is not required. In immunocompromised individuals though, the viral titres are generally lower than neonates and other herpesviruses are often present in the urine. • (f) ELISAs for CMV antigen in the urine - these tests carry low sensitivity as CMV is complexed to ß2-microglobulin in the urine. • (g) Detection of CMV DNA by PCR - the use of PCR in the diagnosis of CMV infection had been widely studied. PCR offers the advantages of being rapid and sensitive. However, its inherent sensitivity poses a problem since latent CMV genomes, which are present in practically all seropositive individuals, may be detected. Therefore, it is critical to adjust the sensitivity of the PCR so that latent genomes are not detected. • (h) CMV antigenaemia test - this test is based upon the detection of pp65, a structural protein expressed on the surface of infected polymorphonyclear leucocytes. The number of infected leucocytes present had been reported to correlate with the severity of infection. The main advantage of this test is that it is very rapid so that a result can be available within the same day. As a result, this test is now widely used especially in the monitoring of transplant recipients
  18. 18. • • • • • II- Serology ; - CMV IgM antibodies are detected in primary infection and lasts 3 - 4 months. It is not detectable in recurrent infection except in immunocompromised patients where it is detectable in about a third of the cases. CMV IgM may be undetectable in primary infection in immunocompromised individuals. Solid phase sandwich or antibody capture ELISAs or RIAs are now in routine use. Interference by rheumatoid factor should be excluded. CMV IgM can be sought for in the cord blood samples from infants who are suspected of being congenitally infected and the titre present is generally related to the outcome. However the best method for diagnosing congenital CMV infection remains virus isolation. • CMV IgG is produced early in primary infection and persists lifelong. The detection of CMV IgG is useful as an "immune status screen" (Seropositive individuals are not protected from reactivation of reinfection). Rising titres of IgG can be used as markers of acute infection. This is particularly useful in diagnosing recurrent infections in normal individuals, and in immunocompromised patients who may not develop a IgM response to primary infection. Various methods are used for detecting CMV IgG including CFT, IFT, latex agglutination, ELISAs and RIAs. • Where possible, serological investigation should be backed by virus culture, especially in the case of immunocompromised patients who may fail to mount an immune response. CMV IgG may also be transferred by blood products which may produce false positive results.
  19. 19. Epidemiology • Cytomegalovirus is endemic in all parts of the world; epidemics are unknown. It is present throughout the year, with no seasonal variation seen in infection rates. • The prevalence of infection varies with socioeconomic status, living conditions, and hygienic practices. Humans are the only known host for cytomegalovirus. Transmission requires close person-to-person contact. Virus may be shed in urine, saliva, semen, breast milk, and cervical secretions and is carried in circulating white blood cells. Oral and respiratory spread are probably the dominant routes of cytomegalovirus transmission. It can also be spread transplacentally, by blood transfusion, by organ transplantation, and by sexual contact. • • • • • • New infections are almost always asymptomatic. After infection, virus is shed from multiple sites. Viral shedding may continue for years, often intermittently, as latent virus becomes reactivated. Thus, exposures to cytomegalovirus are widespread and common.
  20. 20. • Intrauterine infection may produce serious disease in the newborn. • The majority have subclinical but chronic infections; 5–10% have cytomegalic inclusion disease with attendant developmental defects and high mortality. • Congenital infections, whether subclinical or clinically apparent, result in chronic infections, with viral shedding detectable for years. • Many more infants become infected with cytomegalovirus in the first months of life, often from infected breast milk or by nursery spread. Most of these infections are subclinical but are usually chronic, with persistent viral shedding.(acquired) • Cytomegalovirus can be transmitted by blood transfusion. • Seronegative solid organ transplantation recipients are at risk, as a seropositive organ transmits the virus in 60–80% of cases. • Cytomegalovirus infections are markedly increased in immunosuppressed populations; transplant recipients often develop infections, most of which are due to reactivations of their own latent virus
  21. 21. Treatment & Control • Drug treatments of cytomegalovirus infections have shown some encouraging results. Ganciclovir, a nucleoside structurally related to acyclovir, has been used successfully to treat life-threatening cytomegalovirus infections in immunosuppressed patients. • The severity of cytomegalovirus retinitis, esophagitis, and colitis is reduced by ganciclovir. In addition, early treatment with ganciclovir reduces the incidence of cytomegalovirus pneumonia in bone marrow allograft recipients. • Ganciclovir also controls progressive hearing loss in neonates with congenital infections. renal transplant patients. • Specific control measures are not available to prevent cytomegalovirus spread. Isolation of newborns with generalized cytomegalic inclusion disease from other newborns is advisable.
  22. 22. • Screening of transplant donors and recipients for cytomegalovirus antibody may prevent some transmissions of primary cytomegalovirus. • The cytomegalovirus-seronegative transplant recipient population represents a high-risk group for cytomegalovirus infections. Administration of human IgG prepared from plasma pools obtained from healthy persons with high titers of cytomegalovirus antibodies (cytomegalovirus immune globulin) has given discordant results in tests to decrease the incidence of viral infections in transplant recipients. Cytomegalovirus immune globulin is in limited supply. • The use of blood from seronegative donors has been recommended when infants will require multiple transfusions. This approach would eliminate transfusion-acquired cytomegalovirus infections, but it is difficult to implement. • Both live and recombinant cytomegalovirus vaccines are under development