Polymerase chain reaction for diagnosis and identification of distinct variants of Crimean-Congo hemorrhagic fever virus in the United Arab Emirates.

Viral hemorrhagic fever has re-emerged in the United Arab Emirates (UAE) since November 1993. Genomic RNA of Crimean-Congo hemorrhagic virus (C-CHFV) was detected by a newly developed, nested reverse transcriptase polymerase chain reaction (RT-PCR) in the sera of four (25.0%) of 16 suspected cases of viral hemorrhagic fever. The RT-PCR was based on oligonucleotide primers deducted from the small RNA segment encoding the nucleoprotein of the virus. By comparison with a nucleotide sequence of a C-CHFV isolate from a Chinese sheep, a divergence of 10.0-11.8% was detected in the C-CHFV variants causing the UAE outbreak. In the four positive sera, three phylogenetically distinct C-CHFV variants were amplified and confirmed by direct sequencing of the PCR fragments. These C-CHFV sequences were obtained directly from sera of infected humans without prior propagation in cell culture. The RT-PCR allows rapid detection of genomic C-CHFV RNA in clinical specimens and study of the molecular epidemiology of this infection.

Characterization of two rotaviruses differing in their in vitro and in vivo virulence.

The proteins, genomic RNA and disassembly conditions and pathogenesis in vivo for a virulent mouse rotavirus (MRV) and a less virulent bovine rotavirus (BRV) were compared. An obvious difference in the mobility of several genomic RNA segments were observed in one-dimensional gels. Reassortants obtained by replacement of gene 4 in BRV with MRV gene 4 indicated that the dose of the virus used and the clinical outcome in vivo was determined by gene segment 4. Under in vitro conditions, a comparison of the inner capsid proteins by partial proteolytic peptide mapping did not reveal any difference between corresponding proteins. However, this technique did identify differences between the two corresponding outer capsid proteins of these viruses. These differences, in turn, may account for the increased stability of MRV, as compared to BRV, when subjected to calcium-chelating and chaotropic agents and may be one of the mechanisms involved in conferring virulence on the virus. The observed variability between the 84K proteins (VP4) provided a basis for further investigations in order to locate a potential virulence determinant, since it has been observed by us and others that the gene coding for this protein plays a role in determining virulence. A comparison of the carboxypeptidase digests of the MRV and BRV VP4 revealed an area of variability between amino acids 307 and 407, which may represent the site of a virulence determinant.

Donor seropositivity and prednisolone therapy as risk factors for cytomegalovirus infection and disease in cyclosporin-treated renal allograft recipients.

We attempted to assess the importance of blood transfusion, donor seropositivity, and prednisolone therapy as risk factors for cytomegalovirus infection in cyclosporin-treated renal allograft recipients. Primary infection was diagnosed in 27 of 86 patients (31 per cent) and recurrent infection in 27 of 79 patients (34 per cent). Receipt of banked blood from unselected donors after transplantation did not increase the incidence of primary infection in the few transfused patients. Kidney donor seropositivity and maintenance prednisolone in addition to cyclosporin were associated with increases in the incidence of primary or recurrent infection, respectively. Cytomegalovirus infection was clinically mild. Presumed bacterial pneumonias occurred in three patients with recurrent cytomegalovirus infection. The absence of severe cytomegalovirus disease probably reflected the minimal use of prednisolone. Matching of seronegative donors with seronegative recipients seemed unjustifiable in cyclosporin-treated renal transplant patients.

Antiphospholipid antibodies and Mycoplasma pneumoniae infection.

Anticardiolipin antibody levels were measured in 57 patients with Mycoplasma pneumoniae infection and 21 patients with other infections. Significantly more patients in the mycoplasma group had increased IgM and IgG anticardiolipin. Within the mycoplasma group significantly higher titres were found in patients with severe infection (assessed by need for hospital admission) and in patients with cold agglutinins. A tendency for particularly high titres to occur in patients with extra-pulmonary complications was identified.

Effects of the anti-cancer agent etoposide on human herpesvirus 1 replication in vitro.

The anti-human herpesvirus type 1 (herpes simplex virus 1; HSV1) activity of etoposide (VP-16-213, a semi-synthetic derivative of epipodophyllotoxin) was investigated in vitro. Etoposide (but not the proprietary solvent in which the compound is usually formulated) demonstrated a significant antiviral action, probably through an effect on virus replication. Etoposide, at 3 micrograms/ml, induced a 50% reduction of HSV1-plaque formation in Vero cells. These findings are considered in the context of the use of etoposide in an in vivo procedure for the diagnosis of herpes encephalitis through virus-specific scintigraphic brain imaging.

Altered in vitro uptake of the radiolabelled antiviral imaging "probe" E-5-(2-125iodovinyl)-2'-deoxyuridine following administration of acyclovir.

Current developments in the use of radiolabelled antiviral drugs as specific "probes" for virus-infected cells in vivo may allow the specific neuroradiological diagnosis of herpes encephalitis. As "blind therapy" with the antiviral drug acyclovir may precede specific neuroradiological diagnosis, the aim of the present study was to investigate whether or not acyclovir interferes with the uptake of the radioprobe E-5-(2-125Iodovinyl)-2'-deoxyuridine (rIVDU) by virus-infected cells in vitro. Acyclovir treatment (0.1 to 10 micrograms/ml) was shown to increase initial radioprobe uptake by virus-infected cells. However, with continued incubation in the presence of acyclovir, intracellular radioactivity decreased to a level not significantly different from that associated with noninfected cells. A mechanism to explain these results is proposed. It was concluded that concurrent acyclovir therapy could interfere with neuroradiological diagnosis using rIVDU, although this may not occur with all the candidate radioprobes currently under investigation.

Altered in vitro uptake of E-5-2-125iodovinyl-2'-deoxyuridine following administration of the antineoplastic agent etoposide.

The semi-synthetic epipodophyllotoxin derivative, etoposide (VP-16-213), has been shown to inhibit nucleoside uptake in mammalian cells. The present study examined whether etoposide (or the solvent in which it is usually supplied) affected the uptake of the radioiodinated antiviral nucleoside analogue E-5-2-125Iodovinyl-2'-deoxyuridine (125IVDU) by HSV1-infected cells (human herpesvirus 1; herpesvirus simplex type 1). Etoposide was found to significantly reduce 125IVDU sequestration, although some of this effect could be attributed to the solvent. The results are discussed in relation to the use of etoposide in the development of a specific, scintigraphic brain imaging technique to enable early diagnosis of herpes encephalitis.

Penetration of the blood-brain barrier by the antiviral drug (E)-5-(2-iodovinyl)-2'-deoxyuridine in a rat model of herpes encephalitis.

The blood-brain barrier penetration of a radiohalogenated (125I) derivative of the antiviral drug (E)-5-(2-iodovinyl)-2'-deoxyuridine (IVDU) was investigated in a rat model of herpes encephalitis. CNS delivery was assessed by external gamma camera scintigraphic imaging in vivo, a technique which may have general application in evaluating brain specific delivery of drugs, and by autoradiography of cryostat sections of rat brain. Radiohalogenated IVDU was found to be almost totally excluded from the CNS. These findings may be explained in terms of the poor lipid solubility of IVDU (in vitro oil/aqueous salt solution partition coefficient 0.012). Since IVDU appears in CSF after carotid artery administration, our results stress the necessity, in the design of compounds for the treatment of CNS infections, of distinguishing blood-brain, blood-CSF, and CSF-brain drug barriers. The significance of our data both in relation to the development of neuro-radiological diagnosis and antiviral chemotherapy of CNS infection is discussed.

Human papillomavirus (HPV) genotypes associated with laryngeal papilloma.

Biopsy specimens from 14 patients treated for laryngeal papillomatosis were tested for the presence of human papillomavirus (HPV) genome by the technique of DNA-DNA hybridisation. According to the age of initial presentation, cases were subdivided into juvenile (less than 16 years) and adult onset (older than 16 years) groups. Histological investigation confirmed that it was impossible to distinguish the groups on this basis. Molecular virology using both dot blot and Southern transfer techniques showed that 10 cases carried the HPV type 6 genome, three cases HPV type 11, and in one case no HPV DNA was detected. All six adult onset cases carried HPV 6 sequences while the juvenile onset group comprised four HPV 6 and three HPV 11 cases. In the juvenile onset group more females were affected; in the adult onset group more males were affected. Two of the patients shown to have HPV type 11 sequences in their biopsy material were the most resistant to treatment. One of the adult onset cases subsequently developed a squamous cell carcinoma of the larynx in which HPV 6 DNA was detected. As far as we know this is first time that HPV-DNA has been confirmed in laryngeal papilloma undergoing malignant change.

Specific neuro-radiological diagnosis of herpes encephalitis in an animal model.

The potential of utilizing a radio-labelled derivative of the antiviral drug (E)-5-(2-iodovinyl)-2'-deoxyuridine (IVDU) for the specific, non-invasive, in vivo diagnosis of Herpes simplex virus encephalitis (HSVE) was investigated in a rat model of the disease. Following pharmacological disruption of the blood brain barrier radiolabelled IVDU was administered by intra-carotid injection. Brain radioactivity was compared between control and infected animals via gamma camera scintigraphy. After clearance of non-metabolized drug, markedly higher levels of activity were found in infected brain. Post-mortem studies of cryostat sections of brain examined by autoradiography and immunochemical staining showed the radioactivity selectively accumulated in areas of virus infection. These results indicate that radio-labelled derivatives of antiviral drugs may allow the specific neuro-radiological diagnosis of HSVE.

A rat model of herpes encephalitis with special reference to its potential for the development of diagnostic brain imaging.

A rat model of herpes encephalitis using intraocular inoculation of herpes simplex virus strain SC16 was investigated. Virus distribution in the brain was examined by virus isolation and immunocytochemical staining using immuno-gold silver and peroxidase-anti-peroxidase. At 5 days post-infection virus was found in the thalamus, hypothalamus, septum, colliculus, geniculate bodies, the pons, trapezoidium and medulla oblongata, but less frequently, in the cerebellum and occipital lobes. Possible routes of spread of virus and the potential of this model in neuro-radiological scanning procedures are discussed.

Chronic Q fever endocarditis with massive splenomegaly in childhood.

Two children with congenital heart disease developed persistent fever, anemia, and hepatosplenomegaly. Both were shown to have intracardiac vegetations and evidence of infection with Coxiella burnetti. Thus, the same clinical manifestations of Q fever may develop in both children and adults.

Effect of selective vaccination on rubella susceptibility and infection in pregnancy.

The effect of school and adult vaccination on susceptibility to rubella in women of childbearing age was assessed in the Manchester area, where the population attending antenatal clinics is over 40 000 a year. Between 1979 and 1984 the proportion susceptible fell from 6.4% to 2.7%. In 1984, 4.2% of nulliparous women were susceptible compared with 1.4% of women in their second or subsequent pregnancy. Eighty five per cent of pregnant women screened and found to be non-immune were vaccinated post partum before leaving hospital. Requests for prevaccination screening of non-pregnant women increased in response to a national campaign and at the time of local outbreaks of rubella but only two thirds of those found to be nonimmune were subsequently vaccinated. During 1983 and 1984 infection was confirmed in 57 pregnant women--2% of those non-immune. Selective vaccination has reduced susceptibility to rubella in the childbearing population, but it is suggested that mass vaccination of children of both sexes should be added to the existing policy to control circulation of wild rubella virus and reduce the risk of infection to pregnant women who remain susceptible.

Herpes simplex infections in atopic eczema.

One hundred and seventy nine children with atopic eczema were studied prospectively for two and three quarter years; the mean period of observation being 18 months. Ten children had initial infections with herpes simplex. Four children, very ill with a persistently high fever despite intravenous antibiotics and rectal aspirin, continued to produce vesicles and were given intravenous acyclovir. There were 11 recurrences among five patients. In two patients the recurrences were as severe as the initial lesions, and one of these children had IgG2 deficiency. Use of topical corticosteroids preceded the episode of herpes in only three of the 21 episodes. Symptomatic herpes simplex infections are common in children with atopic eczema, and are suggested by the presence of vesicles or by infected eczema which does not respond to antibiotic treatment. Virological investigations are simple and rapid: electron microscopy takes minutes, and cultures are often positive within 24 hours.

Mild forms of herpes encephalitis.

Three cases of herpes encephalitis are described. A definite diagnosis was established and all patients made a good recovery without specific antiviral chemotherapy. These reports are representative of those forms of herpes encephalitis with a good prognosis. It is suggested that the introduction of rapid non-invasive procedures will indicate a higher incidence of herpes encephalitis than is presently accepted. The relevance of repeated episodes of herpes encephalitis with respect to the aetiology of some psychiatric disorders and in the evaluation of antiviral agents is also discussed.

Meningo-encephalitis caused by Coxsackievirus group B type 2. Diagnosis confirmed by measuring intrathecal antibody.

We describe a case of meningo-encephalitis caused by Coxsackievirus group B type 2. Diagnosis was suggested by isolation of virus from faeces and high titres of serum antibody in which Coxsackie virus B2 specific IgM antibody was detected. Evidence of intrathecal production of antibody confirmed the diagnosis.

A simple method for immune analysis of proteins following sodium dodecyl sulphate polyacrylamide gel electrophoresis.

Immune analysis of polypeptides separated by SDS-PAGE provides a technique useful in the characterisation of complex mixtures. We describe a simple procedure involving immune analysis of fractionated PAGE gels. Polypeptides of purified herpesvirus simplex, HSV 'excreted antigen' and cell culture 'control' preparations were separated by SDS-PAGE. Following electrophoresis, protein was eluted from gels, adsorbed to plastic and analysed using an RIA technique. When using a mouse anti-HSV1 serum, 7 peaks of immune reactivity were observed with pure virus and 6 with HSV 'excreted antigen'. Only 1 peak of reactivity was observed with a 'control' antigen.