Reoviridae

Reoviridae

1. Reoviridae By- Sanju Sah St. Xavier’s College, Maitighar , Kathmandu Department of Microbiology

2. Reoviruses , Rotaviruses, Orbiviruses and Coltiviruses 6 hrs

3. Introduction Reo : from R espiratory e nteric o rphan In 1959, Sabin proposed the name reovirus to reflect the fact that viruses of this group had been isolated from the respiratory and enteric tracts and were orphan (reo) viruses without known associated disease. The family Reoviridae is divided into several genera. Four of the genera are able to infect humans and animals: Orthoreovirus, Rotavirus, Coltivirus , and Orbivirus . Other genera infect only the plants, insects, and fish.

4. These genera can be divided into two groups; one group contains viruses with large spikes at the 12 vertices on the particle ( eg , Orthoreovirus ) whereas members of the second group appear more smooth, lacking the large surface projections ( eg , Rotavirus ). The Reoviridae viruses are non-enveloped viruses with double-layered protein capsids. The genome consists of a 10–12 segmented double-stranded RNA with a total genome size of 16–27 kbp depending on the genus Rotaviruses contain 11 genome segments , whereas orthoreoviruses and orbiviruses each possess 10 segments and coltiviruses have 12 segments .

5. The individual RNA segments vary in size from 680 bp (rotavirus) to 3900 bp ( orthoreovirus ). The virus core contains many enzymes essential for transcription and capping of viral RNA. These viruses are unusually resistant to heat, a wide pH (3.0–9.0), and to lipid solvents but are sensitive to 95% ethanol, phenol, and chlorine

6. Important Properties of Reoviruses

7. Diseases associated with reoviruses * Biliary atresia is a rare condition in newborn infants in which the common bile duct between the liver and the small intestine is blocked or absent

8. Structure and composition The virions of Reoviridae family viruses measure 60-80 nm in diameter and possess 2 concentric capsid shells, each of which is icosahedral. No envelope is present. The inner capsids of all genera display sharply defined subunits; the outer capsids of rotaviruses and orbiviruses lack well-defined subunit structures. The genome consists of double-stranded RNA in 10-12 discrete segments, with a total genome size of 16-27 kilobase pair (kbp), depending on the genus.

9. The double-shelled particle is the complete infectious form of the virus. The individual RNA segments vary in size from 680 base pair ( bp ), i.e., rotavirus, to 3900 bp , i.e., orthoreovirus . The virion core contains several enzymes needed for transcription and capping of viral RNA.

10. Structure of family Reoviridae

11. REPLICATION:CYTOPLASMIC Attachement to host receptors probably mediates endocytosis of virus into host cell. Particles are partially uncoated in endolysosomes , but not entirely, and penetrate in the cytoplasm. Early transcription of the dsRNA genome by viral polymerase occurs inside this sub-viral particle (naked core), so that dsRNA is never exposed to the cytoplasm. Transcription from each of the dsRNA segments produces mRNA templates for translation. Viral proteins and genomic RNAs aggregates in cytoplasmic viral factories.

12. (+)RNAs are encapsidated in a sub-viral particle, in which they are transcribed to give RNA (-) molecules with which they become base-paired to produce dsRNA genomes. The capsid is assembled on the sub-viral particle. Mature virions are released presumably following cell death and associated breakdown of host plasma membrane.

13. Viral replication

14. Rotaviruses Rotaviruses are a major cause of diarrheal illness in human infants and young animals, including calves and piglets. Infections in adult humans and animals are also common

15. Morphology Rotavirus shows following features: The virus measures 70 nm in diameter and possesses three-layer icosahedral capsid without an envelope. The sharply defined circular outline of the outer capsid gives the appearance of the rim of a wheel placed on short pokes radiating from a wide arm. This appearance gives the virus the name “ rota ” (Latin word which means wheel). The genome consists of a 11 segmented double-stranded RNA genome .

16. The outer shell is composed of a major glycoprotein with a molecular weight of 34,000 Da and this protein is known as viral protein (VP7). It also consists of a minor trypsin-sensitive protein with a molecular weight of 84,000 Da and this protein is designated as VP4 The minor shell or core protein of the virus core consists of four proteins: VP1, VP2, VP3, and VP6. The virus also consists of six nonstructural proteins

17. Schematic representation of a rotavirus virion. The virus is composed of three protein shells, an outer capsid, an inner capsid, and an internal core, that surround the 11 segments of double-stranded RNA. The outer capsid proteins VP4 and VP7 are neutralization antigens and define the P and G serotypes, respectively. VP6, the inner capsid structural protein, is the subgroup antigen.

18. Rotaviruses: serotypes , groups, and subgroups HRVs belonging to 11 G serotypes have been isolated, but the vast majority have been identified as G1, G2, G3, or G4, and strains belonging to these G types have commonly been designated as serotype 1, 2, 3, or 4, respectively. The severity of illness caused by viruses that belong to these 4 serotypes varies little, if at all. At least 6 different HRV P types have been identified. P type 1a, the most common, is usually associated with G types 1, 3, or 4; whereas, P type 1b is usually associated with G type 2.

19. Rotaviruses are classified into seven groups (from A to G) based on the antigenicity of VP6 and the electrophoretic mobility of the genomic segments. Human disease is caused mostly by group A and occasionally by group B and C rotaviruses

20. Viral replication Rotaviruses replicate in cytoplasm of the host cell . Infection of the cell begins by attachment to the cell surface at the site of beta-adrenergic receptor via VP4 protein and occurs by clathrin-mediated endocytosis Particles are partially uncoated in endolysosomes (loss of the VP4-VP7 outermost layer), and penetrate into the cytoplasm via permeabilization of host endosomal membrane. Early transcription of the dsRNA genome by viral polymerase (RNA dependent RNA polymerase) occurs inside these now double-layered particles (DLPs), so that dsRNA is never exposed to the cytoplasm.

21. The nascent (+)RNAs are extruded into the cytoplasm and serve as template for viral proteins synthesis. The transcribed mRNAs are extruded into the cytoplasm and encode a variety of structural and nonstructural proteins. RNA polymerase is one of those proteins that synthesizes minus strands, which become a part of progeny virus . Progeny cores with replicase activity are produced in cytoplasmic virus factories (also called viroplasms ). This implies synthesis of complementary (-)RNA and initial steps of viral morphogenesis. Late transcription occurs in these progeny cores.

22. At the periphery of virus factories, these core are coated with VP6, forming immature DLPs that bud across the membrane of the endoplasmic reticulum, acquiring a transient lipid membrane which is modified with the ER resident viral glycoproteins NSP4 and VP7; these enveloped particles also contain VP4 . As the particles move towards the interior of the ER cisternae, the transient lipid membrane and the nonstructural protein NSP4 are lost, while the virus surface proteins VP4 and VP7 rearrange to form the outermost virus protein layer, yielding mature infectious triple-layered particles.

23. Mature virions are released presumably following cell death and associated breakdown of host plasma membrane.

24. Pathogenesis and Immunity Rotavirus is transmitted by feco –oral transmission . Rotavirus survives the acidic environment in the stomach and initiates infection in the mucosal cells of the small intestine. After absorption, the viruses replicate in the cytoplasm of the enterocytes and damage their transport mechanism . Non-structural protein 4 (NSP4) of the rotavirus may act as a viral enterotoxin, which causes secretion of fluids by stimulating a signal transduction pathway*. *Signal transduction (also known as cell signaling) is the transmission of molecular signals from a cell's exterior to its interior

25. The toxin induces influx of calcium ion into enterocytes and release of neuronal activators, and alters sodium and glucose absorption. The resulting diarrhea is due to impaired sodium and glucose absorption, as damaged cells on the villi are replaced by non-absorbing immature crypt cells The rotavirus after replicating in the cell causes damage to the cell. The damaged cells are released into the lumen of intestine , releasing large quantities of viruses in the diarrheic stool.

26. It takes around 3–8 weeks for restoring the normal function of the cell. Hence, the rotavirus produces watery diarrhea similar to that seen in cholera

27. Host immunity Rotavirus infection is characterized by the presence of high quantity of immunoglobulin A (IgA) in the intestinal secretions . The IgA plays an important role in conferring the gut immunity against rotavirus. It protects newborns up to the age of 6 months. The serum antibodies usually reduce the severity of the disease, but not necessarily prevent reinfection . Even a small quantity of a virus may cause infection and diarrhea in the absence of specific antibodies in the serum .

28. Clinical Symptoms Rotavirus is a major cause of diarrhea in infants and young children . Diarrhea The incubation period is short being less than 48 hours. The condition manifests commonly as fever, vomiting, diarrhea, and occasionally dehydration. Vomiting is usually of short duration and can occur before or after the onset of diarrhea. The diarrheic stool may be watery, green, or yellow but does not contain mucus.

29. Rotavirus diarrhea is a self-limiting disease, and patients recover completely within 5–10 days without any complications on sequelae. Also, rotavirus has been reported as an agent of traveler’s diarrhea in adults. The virus has also been reported to cause gastroenteritis in adults.

30. Laboratory Diagnosis Specimen Diarrheic stool is the specimen of choice for demonstration of rotavirus and viral antigens . Microscopy Rotavirus can be demonstrated in stool by direct electron microscopy (EM) and by immunoelectron microscopy (IEM )

31. Direct antigen detection Enzyme immunoassay are useful tools to detect rotavirus antigen directly in the stool for diagnosis of diarrheal illness. This is a method used for routine diagnosis of rotavirus diarrhea. Isolation of the virus Rotaviruses are difficult to grow in cell culture, hence are not routinely used for diagnosis of rotavirus diarrhea

32. Treatment No specific antiviral therapy is available for rotavirus infection . The treatment of the condition is mostly supportive. It consists of restoring the fluid loss in dehydrated patients. Oral rehydration fluid using glucose and electrolyte solution is currently preferred than the intravenous fluids to restore the hydration. Intravenous fluids are used only in severely dehydrated children.

33. Prevention A rhesus–human reassortant rotavirus tetravalent vaccine ( RRV-TV; Rota shield) is a being used since 1998 . The vaccine consists of a polyvalent preparation consisting of the VP7 protein of each of the four clinically important serotypes with the attenuation phenotype of the rhesus rotavirus . This has been successful in preventing severe rotavirus diarrhea

34. A pentavalent human-bovine reassortant live-attenuated, oral vaccine ( RotaTeq ) has been developed This vaccine contains five live reassortant rotaviruses Four reassortant rotaviruses express the VP7 protein (G1, G2, G3, or G4) from the human rotavirus parent strain and the attachment protein (P7[5]) from bovine rotavirus parent strain WC3. The fifth reassortant virus expresses the attachment protein (P1A[8 ] from the human rotavirus parent strain and the outer capsid protein G6 from the bovine rotavirus parent strain . RotaTeq is administered in three oral doses at 1- to 2-month intervals beginning at 6 to 12 weeks of age.

35. Orthoreovirus Reoviruses (which also are called orthoreoviruses to avoid confusion with the family Reoviridae ) are nonenveloped viruses. Reovirus particles are composed of an inner protein shell ( ie , core) of a diameter of 60 nm, which is surrounded by an outer protein shell ( ie , outer capsid) that measures 80 nm in diameter. The core is composed of 3 major ( ie , lambda-1, lambda-2, sigma-2) and several minor proteins that surround 10 segments of double-stranded RNA. The virions also contain 3 nonstructural proteins.

36. Reoviruses are ubiquitous, with a very wide host range. Three distinct but related types of reovirus have been recovered from many species and are demonstrable by Nt and HI tests. Reoviruses contain a hemagglutinin for human O or bovine erythrocytes Reoviruses are moderately heat-stable, stable through a wide pH range, and stable in aerosols, particularly when the relative humidity is high . For recovery from clinical material, Reoviruses can be cultured in monkey kidney cells, HeLa cells , etc Because of their widespread prevalence in nature and the ease of infection in laboratory animals, reoviruses have been used widely in pathogenicity studies

37. Human volunteers’ studies have failed to establish a clear cause-and-effect relationship between reoviruses and human ill-ness . So far, reoviruses have been linked with upper respiratory infection , fever, enteritis, and febrile exanthema in children. All three serotypes of the virus have been recovered from healthy children and from children with minor febrile illness, diarrhea, or enteritis . The exact method of transmission of reoviruses is not known . Since viruses are isolated most frequently from the feces, the infection appears to be transmitted by the fecal–oral route.

38. The laboratory diagnosis of the human orthoreovirus infection can be made by: The isolation of the virus and detection of the viral anti-gen and RNA genome in various clinical specimens, such as feces , throat swabs, and nasopharyngeal specimens. The serological tests, such as hemagglutination inhibition, complement fixation, or virus neutralization to demonstrate antibodies. These serological tests are used primarily for epidemiological studies. No specific treatment is available for orthoreovirus infection. No preventive measures have been suggested due to the lack of definitive association of orthoreovirus with human disease.

39. Orbivirus Orbiviruses commonly infect insects, and many can be transmitted by insects to vertebrates. More than 100 serotypes have been identified. Serious animal pathogens include bluetongue virus of sheep and African horse sickness virus. Antibodies are found in many vertebrates, including humans. The genome consists of 10 segments of double-stranded RNA. The replicative cycle is similar to that of reoviruses . In contrast with the general stability of other reoviruses , orbiviruses are inactivated by low pH.

40. Orungo virus, isolated in Africa, has been implicated in an acute illness with myalgias and headache. Lebombo virus is another orbivirus isolated from humans in Africa. Kemerovo virus has been implicated in neurologic infections in central Europe and Russia. Serologic evidence of infection with Lipovnik or Tribec virus has been demonstrated in patients with polyradiculitis in the former Czechoslovakia. Changuinola virus has been isolated from humans in Panama.

41. Coltivirus Coltiviruses resemble the orbiviruses in size and in having 2 capsids. These viruses contain 12 RNA segments. Colorado tick fever (CTF), Salmon River virus (in Idaho), Eyach virus (in Germany and France), isolate S6-14-03 (in California), Banna virus, Beijing virus, and Gansu virus (in China) either are proved or suspected of causing human disease.

42. Colorado tick fever is an acute viral infection transmitted by the bite of wood tick ( Dermacentor andersoni ) caused by Colorado tick fever virus. Colorado fever was so named because the illness was believed to occur predominantly in Colorado and was used to distinguish this clinical illness from that of Rocky Mountain spotted fever caused by Rickettsia species. The causative agent of this fever was recognized as a virus in 1946 Colorado tick fever virus is a double-stranded RNA virus surrounded by two capsids.

43. The virus contains 12 RNA segments . The virus infects and replicates in the bone marrow, lymph nodes, spleen, and liver of rhesus monkey, but without producing any histological abnormalities. The virus has been shown to replicate in erythroid precursor cells without severely damaging them and is present in mature red blood cells . Colorado tick fever occurs almost exclusively in the Western United States and in Canada. The disease is transmitted by the bite of wood tick, D. andersoni . Larval and nymphal stages of D. andersoni ticks usually transmit the virus among all rodents, but only adult ticks transmit the virus to humans.

44. The ticks acquire the infection on feeding an infected viremic host. The infected ticks subsequently transmit the virus through their saliva during act of feeding on a new susceptible host. Squirrels , rabbits , and deer are the natural animal hosts for the virus . The virus generally causes a nonspecific febrile illness. The incubation period is short and varies from 3 to 6 days. The clinical manifestations of the acute condition are characterized by the sudden onset of fever, chills, headache with retro-orbital pain, malaise, nausea, and occasionally vomiting.

45. A rash is generally absent by which Colorado tick fever is differentiated from the Rocky Mountain spotted fever . The fever is typically biphasic (i.e., with two episodes of fever), each of which lasts 2–3 days, separated by a remission of approximately equal duration. In most cases, the febrile period is followed by moderate to marked weakness and malaise. Complications are rare , but may include hepatitis, pericarditis, epididymoorchitis , atypical pneumonitis, encephalitis, and aseptic meningitis . Viruses are present in the erythrocytes during the first 2 weeks of disease.

46. This is followed by a period during which the viruses infect and replicate within the erythropoietic cells. In the infected red blood cells, the viruses can live for the life of the cells, which is nearly 120 days. A single attack of infection usually produces lifelong immunity. No specific antiviral treatment is available for Colorado tick fever . The condition is usually self-limited and can be prevented by avoiding contact with the wood tick