Coronaviruses 24 Muharram 1430 H 20 February 2009

Coronaviruses 24 Muharram 1430 H 20 February 2009 SBM 2044

Coronaviruses n n n Large enveloped RNA; ss positive-sense Cause common cold. Cause of SARS = severe acute respiratory syndrome Replication occurs in cytoplasm, the mature form is budded into ER and golgi. In humans, coronaviruses are limited to upper resp T.

SARS n n n 2003 – outbreak of SARS inc serious resp disease, pneumonia and progressive resp failure. Begun in rural southern China, where people, pigs and domestic fowl live close together and widespread use of wild species for food and traditional medicine – conditions that promote the emergence of new viral strains. Transmitted by close contact with SARS patients or recent travel to area infected with SARS Incubation period ~6 days Symptoms: headache, dizziness, sorethroat, shortness of breath and 10% death. Diagnosis: by serological test, passive haemagglutination – RBC coated with coronavirus are agglutinated by Abcontaining sera. Rx: No vaccine. Precaution and less travelling.

Viruses that invade the CNS Rabies & Prion Diseases

Rabies viruses: Do VAMPIRES exist?

Rabies n n n Rabies viruses invade CNS, causing an acute infection of CNS. Rabies is a rhabdovirus; rod and bullet-shapped particles; membranous envelope with protruding spikes. Ss RNA negative-sense; virions containing RNAdependent RNA polymerase. Natural hosts are all warm-blooded animals inc foxes, coyotes, bats, skunks. Virus is widely distributed in nervous system, saliva, urine, lymph, milk and blood. Recovery is rare except in certain bats. Vampire bats may transmit the virus for months without any clinical symptoms.

Pathogenesis n n Rabies virus multiplies in muscle and connective tissue at the site of inoculation, and then enters peripheral nerves at NMJ and spreads up the nerves to CNS. Multiplication in CNS leads to encephalitis. Then virus spreads to salivary gland other tissues. Highest titres of the virus is detected in submaxillary salivary gland. Rabies virus produces specific eosinophilic cytoplasmic inclusion, the Negri body, in infected nerve cells. These Negri bodies are filled with viral nucleocapsids.

Negri body in rabies-infected nerve cells

Clinical Findings n n Incubation period is 1 -2 months, but could be as short as 1 week. Symptoms: Malaise, anorexia, headache, photophobia. During acute neurologic phase, which lasts 2 -7 days, patients show signs of NS dysfunction, such as nervousness, apprehensions, hallucinations and bizarre behaviour. Rabies should be considered in any case of encephalitis or myelitis of unknown cause. Long incubation period – people may forget a possible exposure incident, no recollection of being bitten by bat.

Treatment n n Treatment is no benefit after the onset of clinical illness. Therefore prompt post-exposure treatment is vital – clean wounds, administration of rabies immunoglobulin and vaccination. Prompt vaccination to depress viral replication and stop virus from invading CNS as well as to lower concentrations of virus. Vaccine also inactivate rabies virus. Rabies Abs are administered either as pre-exposure prophylaxis (for high risk people); or post-exposure prophylaxis. Vaccination also for dogs, and eliminating stray animals

So, do VAMPIRES exist? n Actually human-to-human infection is very rare. The only documented cases involve rabies transmitted by corneal transplants, whereby the cornea was donated from a patient with undiagnosed CNS disease. Recipients died from rabies 50 -80 days later.

Prions n n Some chronic degenerative diseases of CNS are caused by “slow” or chronic, persistent infections by classic viruses eg. transmissible spongiform encephalitis – CJD caused by prions. Other egs are Kuru, scrapie, BSE of cattles, Gerstmann-Sträussker-Scheinker syndrome. Prion = an agent of proteinaceous material devoid of detectable amount of nucleic acid. Prion agent is usually resistant to standard means of inactivation, such as 3. 7% formaldehyde, dry heat, boiling and 50% ethanol. But prion is sensitive to 90% phenol, household bleach and autoclaving (1 hr 121°C). Guanidine thiocyanate is highly effective in decontaminating medical supplies. Incubation periods may be years before clinical manifestations.

Distinguishing hallmarks of prion disease n n n Confined to the nervous system Amyloid plaques may be present Long incubation periods (months decades) precede clinical and chronic illness Always fatal, no known cases of remission or recovery Host shows no inflammatory/immune response – agents are not antigenic. Hence no production of IFN and no effect on B/T lymphocytes.

Amyloid plaques

Scrapie Animals are susceptible to this disease n High titres of prion are found in brain, spinal cord and eye n Development of amyloid plaques in CNS n Pr. P, is a protease-resistant protein 2730 k. Da, purified from scrapie-infected brain. n Pr. Pc Pr. Psc n normal protein larger host-encoded protein

BSE and new v. CJD n n 1986 – Bovine spongiform encephalopathies “Mad cow disease”, emerged in cattle in Great Britain. Was traced to the use of cattle feed that contained contaminated bone meal from scrapieinfected sheep and BSE-infected cattle carcasses. 1993 – peak. 1 million cattle was infected. 1996 – new variant CJD formed, occurs in younger people.

BSE-infected brain cells

Kuru and classic CJD n n These are 2 human spongiform encephalopathies. Kuru only found in eastern highlands of New Guinea, spread by customs surrounding ritual cannibalism of dead relatives. Since the practice has ceased, disease disappeared. CJD develops gradually in humans, progressive dementia, ataxia, myoclonus. Death in 5 -12 months time. Iatrogenic CJD transmitted accidentally by contaminated growth hormone preparations from human cadaver pituitary gland, by corneal transplant, by contaminated surgical instruments and cadaveric human dura mater grafts used for surgical repair of head injury. No suggestions of CJD transmission by blood or blood products.

Human Cancer Viruses

Virus Family Virus Human Cancer Paillomaviridae Human papillomaviruses Genital tumours, squamous cell carcinoma, oropharyngeal carcinoma Herpesviridae Epstein-Barr v Nasopharyngeal carcinoma, African Burkitt’s lymphoma, B cell lymphoma Hepadnaviridae Hepatitis B virus Hepatocellular carcinoma Flaviridae Hepatitis C virus Hepatocellular carcinoma Retroviridae HTL virus Adult T cell leukaemia HIV AIDS-related malignancies

Features of Viral Carcinogenesis 1. Different types of tumour viruses n DNA tumour viruses encode viral oncoproteins that are important for viral replication n RNA tumour viruses carry and RNA-directed polymerase (reverse transcriptase) that constructs a DNA copy of the RNA genome. The DNA copy (provirus) integrates into host DNA and it is from this integrated DNA copy that all proteins of the virus are translated.

Features of Viral Carcinogenesis 2. Multistep Carcinogenesis: n Multistep genetic changes to convert normal cell into a malignant one. n Intermediate stages are designated “immortalisation”, “hyperplasia”, and “preneoplastic”. n It appears that a tumour virus usually acts as a co-factor, provide in only some of the steps required to generate malignant cells ie. viruses are necessary – but not sufficient – for development tumours with a viral aetiology.

Genes Oncogene = gene that causes cancer. In normal cells, known as proto-oncogenes. n The molecular mechanisms responsible for activating a benign proto-oncogene and converting it into a cancer gene vary – but all involved genetic damage. n Tumour suppressor genes = negative regulators of cell growth, also known as the second class of human cancer gene. n

Genes n Prototypes of tumour suppressor genes are Rb (retinoblastoma) and p 53 genes. Inactivation of tumour suppressor genes Rb retinoblastoma p 53 tumour growth Activation of oncogenes Cancer

Retroviruses n n n Retroviruses contain an RNA genome and an RNA-directed DNA polymerase (rev transcrp). Ss RNA, linear, positive-sense. Enveloped. Retroviruses have been isolated virtually from all vertebrate species. Most viruses of a given type are isolated from a single species, though natural infections across species barriers may occur.

DNA Tumour Viruses n n n RNA tumour virus, eg. Retroviruses carry transduced cellular oncogenes that have no role in viral replication. However, DNA tumour viruses carry transforming genes which encode functions required for viral replication and do not have normal homolog in cells. For eg. Human papillomavirus has viral oncoproteins E 6 and E 7 that interacts with cellular p 53 and p. Rb respectively.

Polyomaviruses n n n Ds DNA, circular; the genome enclosed in a nonenveloped. SV 40 virus is from humans and monkeys. Polyomavirus genome expresses “early” and “late” proteins. Early proteins – eg. SV 40 large tumour antigen, is necs for the replication of viral DNA. The late genes encode for the synthesis of coat protein. SV 40 DNA has been detected in human brain tumours, mesotheliomas, bone tumours and lymphomas. Other human polyomaviruses are BK and JC, have been isolated from immunocompromised patients.

Papillomaviruses HPV n n Are highly tropic for epithelial cells of the skin and mucous membrane. Are the cause for skin warts, genital condylomas, laryngeal papillomas, as well as cervical cancer. HPV-16 and HPV-18 are the common cause of cancer. HPV-6 and HPV-11 (which cause benign genital condylomas) are often found in laryngeal papillomas in children. The early proteins (HPV transforming proteins) are synthesised in cancer tissues and able to complex with p. Rb and p 53.

Adenoviruses Commonly infect humans, and cause mild acute respiratory and intestinal infections. n E 1 A early protein complexes with p. Rb; E 1 B and E 40 RF 1 bind with p 53. n No association of adenoviruses with human neoplasms has been found. n

Herpesviruses - HHV n n n EBV causes acute infectious mononucleosis when it infects B lymphocytes B_______ l_______ EBV encodes a viral oncogene protein (LMP 1) that mimics activated growth factor receptor. LMP 1 can transform B lymphocytes. EBNAs (nuclear Ags) are important for immortalisation of B cells, and consistently seen in BL cells. HHV-8 is suspected as being the cause of K_____ s______.

Hepatitis Viruses – HBV & HCV n n n HBV is a risk factor in the development of liver cancer in humans (hepatocellular carcinoma). The mechanism of oncogenesis ? Persistent viral infection leads to necrosis, inflammation cirrhosis - - -> cancer Chronic infection of HCV also leads to hepatocellular carcinoma. Indirect mechanims. . ?

Human cancers Other factors also may attribute to the oncogenesis in humans, besides the tumour viruses. n Lifestyles: diet, smoking. n Availability of vaccines. n Genetics/hereditary n

Human Immunodeficiency Virus

Lentivirus n n The virus contains the 3 genes required for a replicating retrovirus – gag, pol and env. Classification: Lentiviruses have been isolated from many species including at least 26 different African nonhuman primate species. There are 2 distinct types of human AIDS viruses : HIV-1 and HIV-2. The 2 types are distinguished on the basis of genome organisation and phylogenetic relationships with other lentiviruses. Simian immunodeficiency virus (SIV) is a type of monkey lentivirus. The SIVs appear to be nonpathogenic in their host species of origin (chimpanzee etc. ) Humans probably get infected with HIV from direct contact with infected primate blood.

HIV n n Human immunodeficiency virus (HIV) is a retrovirus, a member of the Lentivirus genus. Ss positive-sense RNA, linear; enveloped.

Disinfection and Inactivation HIV is completely inactivated by treatment for 10 mins at room temp with any of the following: 10% household bleach, 50% ethanol, 35% isopropanol, 0. 5% paraform and 0. 3% H 2 O 2. n Extreme p. Hs (1 and 13) inactivate the virus. n

Common Features of HIV n n n n Transmission by exchange of body fluids Virus persists indefinitely in infected hosts High mutation rates Virus infection progresses slowly It may take years for disease to develop The virus receptor is the CD 4 molecule expressed on macrophages and T lymphocytes. The CD 4 molecule has a high affinity for the viral envelope. The second co-receptor is required for fusion of the virus with the cell membrane. The binding of virus with the 2 receptors cause conformational changes in the viral envelope, activating the gp 41 fusion peptide and triggering membrane fusion. Chemokine receptors (CCR 5 on mØ and CXCR 4 on lymphocytes) serve as HIV-1 second receptors. These chemokine receptors used by HIV are found on lymphocytes, macrophages, thymocytes as well as neurons and cells in colon and cervix.

CD 4 molecule of T cells n n n Is the major receptor for HIV Number of molecules depleted during HIV infection. The consequences of CD 4 T cell dysfunction are devastating because the CD 4 T lymphocyte plays a critical role in the human response, which inc activation of macrophages, induction of cytotoxic T cells, NK cells and B cells.

n n Individuals who possess homozygous deletions of CCR 5 may be protected from infection by HIV-1; mutations in the CCR 5 gene promoter appear to delay disease progression. A dendritic cell-specific lectin, DC-SIGN, appears to bind HIV-1 but not to mediate cell entry. Rather, it may facilitate transport of HIV by dendritic cells to lymphoid organs and enhance infection of T cells.

Replication of HIV n n HIV binds to CD 4 cell surface molecules, entry into the cell also requires binding to co-receptors. CXCR 4 and CCR 5). This step can be inhibited by fusion/entry inhibitors. HIV is uncoated inside the cell and reverse transcriptase copies genomic RNA into DNA, making errors at a frequence of about one per replication cycle. Reverse transcriptase inhibitors were the first class of HIV inhibitors to be used as drugs. Viral DNA can integrate into DNA and become a part of the cellular genome. This step makes the infection irreversible, and may mean that eliminating the virus from an infected individual is not possible. Integrase inhibitors are designed to block this step of infection. The virus uses cellular machinery to synthesize viral proteins. Several of these are long amino acid chains which must be cleaved by a specific viral protease before new viral particles can become active. Protease inhibitors block viral maturation at this step.

Course of HIV infections n n n Following primary infection, it takes 8 -12 weeks for the viraemia. Virus is widely disseminated throughout the body during this time, and the lymphoid organs become seeded. An acute mononucleosis-like syndrome develops 3 -6 weeks after primary infection, during which a significant drop in circulating CD 4 T cells is also detected. An immune response occurs 1 week to 3 months after infection, however this was not enough to clear the infection completely, and the HIVinfected cells persist in the lymph nodes.

Course of HIV Infection n n The period of clinical latency may last for as long as 10 years. During this time, there is a high level of ongoing viral replication. There is a high turnover rates of HIV, and rapid viral proliferation and high mutation rates. Eventually, the patient will develop constitutional symptoms and clinically apparent disease eg. opportunistic infections or neoplasms. HIV found in patients with late-stage disease is usually much more virulent and cytopathic than strains recovered early in infection.

Clinical Findings n n AIDS is characterised by pronounced suppression of the immune system and development of a wide variety of severe opportunistic infections or unusual neoplasms (Kaposi’s sarcoma). Symptoms: weight loss, candidiasis, hairy leukoplakia. Disease symptoms in GIT from the oesophagus to the colon are a major cause of debility. With no treatment, the interval between primary infection with HIV and the first appearance of clinical disease is usually long in adults; 8 -10 years.

Plasma viral load n n n The amount of HIV in the blood (viral load) is of significant prognostic value. Apparently a single measurement of plasma viral load about 6 mths after infection is able to predict the subsequent risk of development of AIDS in men several years later. Difference in gender – women may be less predictive. Plasma viral load is best at predicting long-term clinical outcome. CD 4 lymphocyte counts are best predictor of short-term risk of developing an opportunistic disease. Plasma viral load is critical at assessing the effectiveness of antiretroviral therapy.

Paediatric AIDS n n n Neonates acquired the HIV from mothers. Clinical symptoms appear at 2 years of age. Death follows 2 years later. Devastating for the neonates since immune system has not developed at the time of primary infection. Symptoms: pneumonia, oral candidiasis. Untreated children will have very poor prognosis. A high rate of disease progression occurs in the first few years of life. Initially, low viral RNA load at birth, which then rapidly rise within the first 2 mths of life, followed by a slow decline until the age of 24 mth – suggesting that the immature immune system has difficulty containing the infection.

Other Associated Diseases n n n A) Neurological disease – in 40 -90% patients. Severe symptoms inc aseptic meningitis, dementia, toxoplasmosis. Paediatric AIDS patients display seizures, loss of behavioral developmnt and attention deficit disorder. B) Opportunistic infections – infections by agents rarely cause a problem. Infections inc Toxoplasma, Candida albicans, Mycobacterium tuberculosis, Listeria monocytogenes, HSV, VZV. C) Cancer is another consequence of immune suppression. AIDS-associated cancers inc non. Hodgkin’s lymphoma (Burkitt’s lymphoma), Kaposi’s sarcoma,

Immunity n n n HIV-infected persons develop both humoral and cellmediated responses. Most individuals make low level of neutralising Abs against HIV, directed against envelope gp. The envelope gp has great variability which allows resistant virus to escape recognition by the host. Cellular responses develop against HIV proteins. CTLs recognise env, pol and gag gene products. NK cells act against HIV-1 gp 120. It is not clear which host responses are crucial against HIV infection.

Epidemiology n n AIDS was first recognised in 1981. By 2002, WHO estimated that more than 24 million people worldwide had died of AIDS. The WHO estimates that of the 5 million new HIV infections each year, 90% are occurring in developing countries. Based on 2002 data, sub-Saharan Africa had the highest number of HIV infections. High prevalence – as many as 1 of every 3 adults was infected.

Routes of Transmission n n High titres of HIV are found in blood and semen. HIV is transmitted sexually. The presence of other sexually transmitted diseases such as syphilis, gonorrhoea or HSV-2 increases the risk of sexual HIV transmission because the inflammation and sores facilitate the transfer of HIV across mucosal barriers. Blood transfusions Injection drug use Mother-to-infant in utero, during birth process, or through breast-feeding (common, highest way of infection in Africa) Health care workers may accidentally get infected following a needlestick with contaminated blood, though the risk is very low 0. 3%.

Prevention & Treatment n n n Antiviral drugs: nucleoside and non-nucleoside inhibitors and protease inhibitor. Highly active antiretroviral therapy (HAART) – to suppress viral replication, decrease viral load in lymphoid tissues and to allow recovery against opportunistic infections. However, HAART has failed. Monotherapy usually results in the rapid emergence of drug-resistant mutants of HIV. Triple-drug therapy has proven effective. Current drug-therapy may have toxic side effects.

Prevention & Treatment n n Zidovudine (azidothymidine, AZT) can reduce the HIV transmission from mother to infant. Vaccine development is difficult because HIV mutates (envelope antigens) rapidly, HIV is not expressed in all cells that are infected and is not completely cleared by the host immune system. Vaccine using recombinant viral proteins (envelope gp) are likely candidates. Gene therapy to genetically alter target cells to make them resistant to HIV.

Control Measures n n n Maintain an ad-Deen healthy lifestyle. All donor blood should be screened for Ab. Educating HIV-infected individuals. Careful when dealing with contaminated syringes, needles and other equipment. Health care professionals should take precautions when performing invasive procedures to HIV-infected patients.