Table of Contents:

• Taxonomy Information
• Lifecycle Information
• Genome Information
• Biosafety Information
• Culturing Information
• Epidemiology Information
• Diagnostic Tests Information
• Infected Hosts Information
• Phinet: Pathogen-Host Interaction Network
• Lab Animal Pathobiology & Management
• References
• Data Provenance and Curators

1. Species:
   1. Rift Valley fever virus :
      1. Common Name: Rift Valley fever virus
      2. GenBank Taxonomy No.: 11588
      3. Description: Rift Valley Fever (RVF) is an arthropod-borne disease primarily causing epizootics of abortion and high mortality in domestic animals during which humans become infected(Meegan and Bailey, 1989). The disease is caused by the RVF virus, a member of the genus Phlebovirus in the family Bunyaviridae. The disease was first reported among livestock by veterinary officers in Kenya in the early 1900s(Website 2).
      4. Variant(s):
         • Rift valley fever virus (STRAIN ZH-548 M12) (Website 3):
           • Common Name: Rift valley fever virus (STRAIN ZH-548 M12)
           • GenBank Taxonomy No.: 11589
           • Parents: Rift Valley fever virus
           • Description: Strain ZH 548 was isolated in Egypt in 1977 from an infected human(Sall et al., 1998). The MP12 attenuated strain of Rift Valley fever virus was obtained by 12 serial passages of a virulent isolate ZH548 in the presence of 5-fluorouracil(Vialat et al., 1997).

1. Rift Valley Fever Virus Information
   1. Stage Information:
      1. Virion:
         • Size: Mature particles, liberated by the disintegration of Vero cells, contained ribosome-like structures within the nucleocapsid, which was surrounded by a typical unit membrane through which were inserted some 350-375 surface spikes whose inner ends were incorporated into the nucleocapsid structure. In the negatively stained material, the overall diameter of the virion was 90-110 nm; the spikes were 10-18 nm in length and 5 nm in diameter.
         • Shape: Viruses of the family Bunyaviridae have similar morphological features (Gonzalez-Scarano and Nathanson, 1996). The viral particles are spherical or pleomorphic, depending on the method used for fixation.

1. Genome of Rift Valley fever virus
   1. Description: Like all the phleboviruses, RVF virus possesses a tripartite genome consisting of negative sense single-strand RNA segments designated L (large), M (medium) and S (small). The L segment codes for the L viral polymerase. The M segment codes for the precursor to the envelope glycoproteins G1 and G2 and the non structural proteins 14 and 78 kDa. The S segment codes for the nucleocapsid protein N and the non structural protein NSs in an ambisense strategy(Vialat et al., 1997).
   2. S segment(Website 11)
      1. GenBank Accession Number: NC_002045
      2. Size: 1690(Website 11).
      3. Gene Count: 2 genes. The S segment codes for the nucleocapsid protein N and the non structural protein NSs in an ambisense strategy(Vialat et al., 1997).
      4. Description: The gene organization and expression of the S segment are the major features that distinguish the Phlebovirus genus from bunyaviruses, hantaviruses, and nairoviruses, but are similar to those topoviruses(Giorgi, 1996). The sequences and coding strategies of the S RNAs of two viruses, Toscana (TOS) and the M12 derivative of Rift Valley fever ZH-548 (RVF, Phlebovirus genus, Bunyaviridae) have been determined from cDNA clones and compared to the previously published sequences of Punta Toro (PT), Sandfly fever Sicilian (SFS), and Uukuniemi (UUK) viruses. All five viruses exhibit an ambisense coding strategy for their small (S) RNA species, i.e., one gene product (the NSs protein) is encoded in the 5' half of the viral RNA, a second (the N protein) is encoded in the sequence complementary to the 3' half(Giorgi et al., 1991).
   3. M Segment(Website 12)
      1. GenBank Accession Number: NC_002044 M25276 M11157
      2. Size: 3884 or 3885 bp(Website 12).
      3. Gene Count: 2 genes. The M segment codes for the precursor to the envelope glycoproteins G1 and G2 and the non structural proteins 14 and 78 kDa(Vialat et al., 1997).
      4. Description: The sequence data show that all viruses possess a single large ORF extending the length of the M segment in the viral complementary RNA. This ORF codes for a polypeptide precursor of the viral glycoproteins G1 and G2(Giorgi, 1996).
   4. L Segment(Website 13)
      1. GenBank Accession Number: NC_002043
      2. Size: 6606(Website 13).
      3. Gene Count: 1 gene. The L segment codes for the L viral polymerase(Vialat et al., 1997).
      4. Description: The L segment codes for the L viral polymerase(Vialat et al., 1997).

1. Biosafety information for Rift Valley fever virus
   1. Level: SALS recommends that work with this agent be conducted only in Biosafety Level 3 facilities which provide for HEPA filtration of all exhaust air prior to discharge from the laboratory(Website 17).
   2. Precautions: Clearly, RVF presents a hazard to all laboratory workers engaged in its study, and appropriate precautions should be taken to prevent infection. These include wearing protective clothing including, at the minimum, goggles, mask, waterproof apron, and gloves. The handling of the virus should be carried out under a hood so designed that the air flows away from the worker to the exhaust, and on the way passes through a battery of ultraviolet lights and a heated chimney. Laminar-flow hoods which achieve the same purpose are standard equipment in most laboratories(Gear, 1988).

1. Cell Culture Infomation :
   1. Description: Rift Valley fever virus replicated readily in most common cell cultures; the virus is cytopathic and forms plaques(Gonzales-Scarano and Nathanson, 1996). Eleven human sera from known cases of Rift Valley fever (RVF) were obtained from the 1987 epidemic in Mauritania and served as the source of virus for these studies. Sera were inoculated directly into cell cultures (Vero, C6/36 and DBS-FRhL-2) and animals (ICR suckling mice, Lak:LVG(SYR) hamsters and WF rats) concurrently. The cell lines provided a quick method to propagate, quantitate and identify these specimens without prior adaptation. The isolates were highly virulent for suckling mice and hamsters, but not for WF rats, even after cell culture passage(Anderson et al., 1989). Vero cells are routinely used at the USAMRIID laboratory to produce RVF viral pools and titrate strains from various regions of Africa(Anderson et al., 1989). Although isolation and titration of RVFV in suckling mice has been considered the traditional and most sensitive method, standard laboratory cell lines would appear to be a more practical and efficient alternative. In this study, cell lines allowed rapid titration of clinical specimens by an easily read plaque assay, produced virus seed stocks in certified cells acceptable for vaccine production and allowed a rapid preliminary viral identification by indirect immunofluorescence. This was possible with clinical specimens without prior adaptation of the viral isolate in tissue culture or animal systems(Anderson et al., 1989).
   2. Medium: The C6/36 subclone of Aedes albopictus cells was grown at 25 degrees C in HMEM medium supplemented with 5% fetal bovine sera and antibiotics. DBS-FRhL-2 and Vero cells were maintained in EMEN supplemented with non-essential amino acids, glutamine, antibiotics, and 10% heat inactivated fetal bovine serum(Anderson et al., 1989).
   3. Note: Diagnosis by viral cultivation and identification for the VHF-causing agents requires 3 to 10 days for most (longer for the hantaviruses); and, with the exception of dengue, specialized microbiologic containment is required for safe handling of these viruses(Website 14). For isolation of VHF viruses from fatal cases, post-mortem tissues are homogenized to a concentration of 10 to 20% in sterile buffer containing antibiotics and clarified by centrifugation. Stool suspensions and urine are also centrifuged at 10,000 x g for 30 min to remove gross bacterial contamination. Attempts to isolate virus from clinical specimens should proceed using laboratory animals or cell cultures. In all cases cell culture should be used in conjunction with animal inoculation for virus isolation. Inoculated animals are examined daily for illness or death and guinea pig rectal temperatures recorded. Both CCHF and Rift Valley fever viruses produce fatal illness in suckling mice(Shepherd, 1988).

1. Outbreak Locations:
   1. RVF is generally found in regions of eastern and southern Africa where sheep and cattle are raised, but the virus also exists in most countries of sub-Saharan Africa and in Madagascar. In September 2000, a RVF outbreak was reported in Saudi Arabia and subsequently Yemen. These cases represent the first Rift Valley fever cases identified outside Africa(Website 2).
2. Transmission Information:
   1. From: Mosquito(Website 2). , To: Human(Website 2). , With Destination:Human(Website 2). (Website 2)
      Mechanism: Humans can get RVF as a result of bites from mosquitoes and possibly other bloodsucking insects that serve as vectors(Website 2).
   2. From: Cattle(Website 2). , To: Human(Website 2). , With Destination:Human(Website 2). (Website 2)
      Mechanism: Humans can also get the disease if they are exposed to either the blood or other body fluids of infected animals. This exposure can result from the slaughtering or handling of infected animals or by touching contaminated meat during the preparation of food(Website 2).
   3. From: Sheep(Website 2). , To: Human(Website 2). , With Destination:Human(Website 2). (Website 2)
      Mechanism: Humans can also get the disease if they are exposed to either the blood or other body fluids of infected animals. This exposure can result from the slaughtering or handling of infected animals or by touching contaminated meat during the preparation of food(Website 2).
   4. From: Goat(Website 2). , To: Human(Website 2). , With Destination:Human(Website 2). (Website 2)
      Mechanism: Humans can also get the disease if they are exposed to either the blood or other body fluids of infected animals. This exposure can result from the slaughtering or handling of infected animals or by touching contaminated meat during the preparation of food(Website 2).
   5. From: Mosquito(Lupi and Tyring, 2003). , To: Mosquito(Lupi and Tyring, 2003). , With Destination:Mosquito(Lupi and Tyring, 2003). (Lupi and Tyring, 2003)
      Mechanism: The interepizootic survival of RVF virus is believed to depend on tranovarian transmission of virus in floodwater Aedes mosquitoes. Virus can persist in mosquito eggs until the next period of heavy rainfall, when they hatch and yield mosquitoes infected with the RVF virus(Lupi and Tyring, 2003).
3. Environmental Reservoir:
   1. Currently no environmental reservoir information is available.
4. Intentional Releases:
   1. Intentional Release Information:
      1. Emergency Contact: If clinicians feel that VHF is a likely diagnosis, they should take two immediate steps: 1) isolate the patient, and 2) notify local and state health departments and CDC(MMWR, 1988). Report incidents to state health departments and the CDC (telephone {404} 639-1511; from 4:30 p.m. to 8 a.m., telephone {404} 639-2888). Information on investigating and managing patients with suspected viral hemorrhagic fever, collecting and shipping diagnostic specimens, and instituting control measures is available on request from the following persons at Centers for Disease Control (CDC) in Atlanta, Georgia; for all telephone numbers, dial 404-639 + extension: Epidemic Intelligence Service (EIS) Officer, Special Pathogens Branch, Division of Viral Diseases, Center for Infectious Diseases (ext. 1344); Chief, Special Pathogens Branch, Division of Viral Diseases, Center for Infectious Diseases: Joseph B. McCormick, M.D. (ext. 3308); Senior Medical Officer, Special Pathogens Branch, Division of Viral Diseases, Center for Infectious Diseases: Susan P. Fisher-Hoch, M.D. (ext. 3308); Director, Division of Viral Diseases, Center for Infectious Diseases (ext. 3574). After regular office hours and on weekends, the persons named above may be contacted through the CDC duty officer (ext. 2888)(MMWR, 1988).
      2. Delivery Mechanism: The VHF agents are all highly infectious via the aerosol route, and most are quite stable as respirable aerosols. This means that they satisfy at least one criterion for being weaponized, and some clearly have the potential to be biological warfare threats. Most of these agents replicate in cell culture to concentrations sufficiently high to produce a small terrorist weapon, one suitable for introducing lethal doses of virus into the air intake of an airplane or office building. Some replicate to even higher concentrations, with obvious potential ramifications. Since the VHF agents cause serious diseases with high morbidity and mortality, their existence as endemic disease threats and as potential biological warfare weapons suggests a formidable potential impact on unit readiness. Further, returning troops may well be carrying exotic viral diseases to which the civilian population is not immune, a major public health concern(Website 14).
      3. Containment: Patients with VHF syndrome generally have significant quantities of virus in their blood, and perhaps in other secretions as well (with the exceptions of dengue and classic hantaviral disease). Well-documented secondary infections among contacts and medical personnel not parenterally exposed have occurred. Thus, caution should be exercised in evaluating and treating patients with suspected VHF syndrome. Over-reaction on the part of medical personnel is inappropriate and detrimental to both patient and staff, but it is prudent to provide isolation measures as rigorous as feasible. At a minimum, these should include the following: stringent barrier nursing; mask, gown, glove, and needle precautions; hazard-labeling of specimens submitted to the clinical laboratory; restricted access to the patient; and autoclaving or liberal disinfection of contaminated materials, using hypochlorite or phenolic disinfectants. For more intensive care, however, increased precautions are advisable. Members of the patient care team should be limited to a small number of selected, trained individuals, and special care should be directed toward eliminating all parenteral exposures. Use of endoscopy, respirators, arterial catheters, routine blood sampling, and extensive laboratory analysis increase opportunities for aerosol dissemination of infectious blood and body fluids. For medical personnel, the wearing of flexible plastic hoods equipped with battery-powered blowers provides excellent protection of the mucous membranes and airways(Website 14).

1. Organism Detection Test:
   1. Electron Microscopy :
      1. Description: RVF virus attained an extracellular titer of at least 3.6 logs/ml 4 hours post infection in CV-1, Vero and BHK-21 cells. At 22 hours post infection, a peak titer of 7.7 logs/ml was reached in CV-1 cells, where 50% of the cells showed cytopathic effect. The same degree of cytopathic effect was only observed 45 hour post infection in the other cell lines tested. At 22 hours post infection, RVF viral antigens were detected by indirect immunofluorescence in the three culture systems; however, the degree of fluorescence and the number of fluorescent cells were much greater in CV-1 than in either Vero or BHK-21 cells. Virus particles were detected by EM 22 hours post infection in CV-1 cells, but in Vero and BHK-21 cell only at 45 hours post infection(Mekki and Van Der Groen, 1981). RVF virions have been visualized in post-mortem liver by electron microscopy(Shepherd, 1988).
   2. Electron Microscopy and Immunofluorescence :
      1. Description: RVF virus attained an extracellular titer of at least 3.6 logs/ml 4 hours post infection in CV-1, Vero and BHK-21 cells. At 22 hours post infection, a peak titer of 7.7 logs/ml was reached in CV-1 cells, where 50% of the cells showed cytopathic effect. The same degree of cytopathic effect was only observed 45 hour post infection in the other cell lines tested. At 22 hours post infection, RVF viral antigens were detected by indirect immunofluorescence in the three culture systems; however, the degree of fluorescence and the number of fluorescent cells were much greater in CV-1 than in either Vero or BHK-21 cells. Virus particles were detected by EM 22 hours post infection in CV-1 cells, but in Vero and BHK-21 cell only at 45 hours post infection(Mekki and Van Der Groen, 1981). Specific identification of RVF antigen has been achieved in post-mortem human liver by immunodiffusion and in animal tissues by immunofluorescence(Shepherd, 1988).
   3. Rhesus monkeys inoculated with Rift Valley fever :
      1. Description: Rhesus monkeys inoculated with Rift Valley fever (RVF) virus provide a model in which serial observations of serum viral antigen and antibodies can be made. In 9 non-fatal and 3 fatal infections, either antigen or IgM enzyme-linked immunosorbent assay (ELISA) antibodies were detected in every serum sample during the acute phase. Furthermore, viral nucleic acid could be detected by filter hybridization in most samples taken on days 1 to 3. Circulation of significant quantities of viral RNA provides an additional approach to the diagnosis and study of RVF(Morrill et al., 1989).
2. Immunoassay Test:
   1. Haemagglutination :
      1. Description: A total of 76 sera from 21 individuals immunized with RVF vaccine were tested by plaque reduction neutralization test (PRNT) and ELISA IgG. Of the 76 sera, 70 were also tested by the hemagglutination inhibition (HI) and complement fixation (CF) tests. Sera from day 0 (n equals 21) and days 6 to 8 (n equals 23) were found to be negative by all methods (ELISA IgG, PRNT, HI, and CF). When bled on days 32 to 34 all had developed antibodies by PRNT(Niklasson et al., 1984). The ELISA IgG was almost as sensitive as the PRNT, missing only one positive serum with a PRNT titer of 1:80. Of the 32 sera with PRNT titers of greater than or equal to 1:10, 26 were also tested by the HI and CF tests. HI was less sensitive than PRNT, detecting only 4 positive sera out of 11 sera with PRNT titers of 1:160. When tested by CF, all 26 sera were found to be negative, except one which had a CF titer of 4 and a PRNT titer of 1:640. All sera with negative PRNT results were negative in other tests(Niklasson et al., 1984). The HI and IFA (IgG and IgM) tests are the most rapid and commonly used tests for serological diagnosis of RVF infection, antibodies generally appearing in the serum 4 to 10 days after the onset of symptoms. The reversed passive hemagglutination and inhibition (RPHI) test appears to be similar in most respects to the HI(Shepherd, 1988).
      2. False Negative: HI was less sensitive than PRNT, detecting only 4 positive sera out of 11 sera with PRNT titers of 1:160(Niklasson et al., 1984).
   2. Complement Fixation :
      1. Description: A total of 76 sera from 21 individuals immunized with RVF vaccine were tested by plaque reduction neutralization test (PRNT) and ELISA IgG. Of the 76 sera, 70 were also tested by the hemagglutination inhibition (HI) and complement fixation (CF) tests. Sera from day 0 (n equals 21) and days 6 to 8 (n equals 23) were found to be negative by all methods (ELISA IgG, PRNT, HI, and CF). When bled on days 32 to 34 all had developed antibodies by PRNT(Niklasson et al., 1984). The ELISA IgG was almost as sensitive as the PRNT, missing only one positive serum with a PRNT titer of 1:80. Of the 32 sera with PRNT titers of greater than or equal to 1:10, 26 were also tested by the HI and CF tests. HI was less sensitive than PRNT, detecting only 4 positive sera out of 11 sera with PRNT titers of 1:160. When tested by CF, all 26 sera were found to be negative, except one which had a CF titer of 4 and a PRNT titer of 1:640. All sera with negative PRNT results were negative in other tests(Niklasson et al., 1984).
      2. False Negative: When tested by CF, all 26 sera were found to be negative, except one which had a CF titer of 4 and a PRNT titer of 1:640(Niklasson et al., 1984).
   3. Sandwich and Capture ELISA :
      1. Description: We report on the development and validation of sandwich and capture ELISAs (both based on inactivated antigen) for detection of IgG and IgM antibody to Rift Valley fever virus in bovine, caprine and ovine sera. Compared to virus neutralisation and haemagglutination-inhibition tests, the IgG sandwich ELISA was more sensitive in detection of the earliest immunological responses to infection or vaccination with Rift Valley fever virus. Its sensitivity and specificity derived from field data sets ranged in different ruminant species from 99.05 to 100% and from 99.1 to 99.9%, respectively. The specificity of IgM-capture ELISA varied between different species from 97.4 to 99.4%; its sensitivity was 100% in sheep tested 5-42 days post-infection. Our results in field-collected, experimental and post-vaccination sera demonstrate that these assays will be useful for epidemiological surveillance and control programmes, import/export veterinary certification, early diagnosis of infection, and for monitoring of immune response in vaccinated animals. As highly accurate and safe tests, they have the potential to replace traditional diagnostic methods, which pose biohazard risks limiting their use outside of endemic areas to high containment facilities(Paweska et al., 2003).
   4. ELISA :
      1. Description: An ascitic fluid containing a mixture of mouse monoclonal antibodies, or a control of normal mouse ascitic fluid, were diluted 1/2,000 in PBS and coated to alternate rows of the plate and incubated for 14-16 hours at 4 degrees C. After incubation, then washing, the mixture of monoclonal anti-RVFV antibodies which had been labeled with biotin was diluted 1/500 and added to all wells. Following incubation and washing, peroxidase-conjugated avidin diluted 1/2,000 was added. The plates were incubated, then washed, and substrate was added and the OD read after 60 min incubation at 37 degrees C(Meegan et al., 1989).
      2. False Negative: Eighty-two sera were positive for isolation by at least one of the three standard virus isolation techniques. When tested in the antigen detection ELISA which employed biotin-labeled monoclonal antibodies, 24 of these 82 sera (29.3%) were positive (Meegan et al., 1989). Considering the high viremia reported for patients infected with RVFV, the ELISA sensitivity of 29.3% appears low. It appears that titers of infectious virus generally must exceed 10 to the third PFU/ml of sera before the ELISA could detect antigen(Meegan et al., 1989).
   5. PRNT and ELISA :
      1. Description: A total of 76 sera from 21 individuals immunized with RVF vaccine were tested by plaque reduction neutralization test (PRNT) and ELISA IgG. Of the 76 sera, 70 were also tested by the hemagglutination inhibition (HI) and complement fixation (CF) tests. Sera from day 0 (n equals 21) and days 6 to 8 (n equals 23) were found to be negative by all methods (ELISA IgG, PRNT, HI, and CF). When bled on days 32 to 34 all had developed antibodies by PRNT(Niklasson et al., 1984). The ELISA IgG was almost as sensitive as the PRNT, missing only one positive serum with a PRNT titer of 1:80. Of the 32 sera with PRNT titers of greater than or equal to 1:10, 26 were also tested by the HI and CF tests. HI was less sensitive than PRNT, detecting only 4 positive sera out of 11 sera with PRNT titers of 1:160. When tested by CF, all 26 sera were found to be negative, except one which had a CF titer of 4 and a PRNT titer of 1:640. All sera with negative PRNT results were negative in other tests(Niklasson et al., 1984).
      2. False Negative: The ELISA IgG was almost as sensitive as the PRNT, missing only one positive serum with a PRNT titer of 1:80(Niklasson et al., 1984). All sera with negative PRNT results were negative in other tests(Niklasson et al., 1984).
3. Nucleic Acid Detection Test: