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Sequences of a new herpesvirus with homology to gammaherpesvirinae were recently identified in AIDS-associated Kaposi's sarcoma (KS). Subsequently this novel virus, called KS-associated virus (KSHV) or human herpesvirus (HHV) 8 was detected in classical KS and AIDS-associated body cavity based lymphomas by polymerase chain reaction. In this report major and minor capsid proteins of HHV-8 were molecularly cloned and produced as recombinant proteins in Escherichia coli. Sera from 69 HIV-1 infected patients with KS, 30 HIV-1 infected patients without KS and 106 control individuals were tested by enzyme-linked immunosorbent assay for anti-HHV-8 capsid IgM and IgG antibodies. Sera from four patients were tested over periods ranging from 18 months to 6 years. IgG antibodies directed against HHV-8 capsid antigens were detected in patients with AIDS-associated KS and in some AIDS patients without KS. Seroconversion with IgM and IgG antibodies directed against HHV-8 capsid proteins occurred more than 1 year prior to diagnosis of KS. In a considerable portion of KS patients no IgM or IgG antibodies against HHV-8 capsid proteins were detected. In these patients there was an inverse relationship between antibodies against HHV-8orf26 and the CD4/CD8 ratio, suggesting that the inconsistency of anti-HHV-8orf26 antibodies is due at least partly to an impaired immune response. No reactivity against HHV-8 capsid antigens was detected in the vast majority of sera from HIV-negative control individuals. Our findings indicate that a specific humoral immune response against capsid proteins is raised in HHV-8 infected individuals, and that anti-capsid antibodies can be used to diagnose HHV-8 infection. The correlation between occurrence of anti-HHV-8 antibodies and KS supports the hypothesis of a causative role of HHV-8.

Herpes simplex virus encephalitis is a severe sporadic encephalitis in man with high mortality and morbidity. A critical step in the establishment of therapy is early diagnosis. Magnetic resonance imaging is a noninvasive, accurate diagnostic test for the detection of central nervous system disease. In an effort to monitor morphological changes in vivo we present a new diagnostic neuroimaging model of experimental herpes simplex virus encephalitis. A mouse model of herpes simplex virus encephalitis was used. 40 SJL mice were intranasally inoculated with an infectious dose of wild-type strain HSV-I F. Morphological abnormalities were studied by cranial magnetic resonance imaging (MRI). These findings were correlated with sequential neuropathological studies. 95% of animals developed cerebral abnormalities on MRI. resembling human HSVE. Areas of increased signal intensity on T2-weighted sequences and focal pathological contrast enhancement were mostly found in the frontal and temporal lobes and thalamic and cerebellar regions. All animals with MRI abnormalities had neuropathological signs of neuronal degeneration and reactive astrocytosis in corresponding regions. The described monitoring system offers a new approach for studies on neurovirulence and therapeutic strategies.

The gene encoding the DNA (cytosine-5) methyltransferase (m5C-MTase) of lymphocystis disease virus (flounder isolate, LCDV-1) has been identified by polymerase chain reaction (PCR) using oligonucleotide primers synthesized corresponding to different regions of the m5C-MTase gene of frog virus 3 (FV3). A DNA fragment of 487 bp was amplified using oligonucleotide primers L3 and R4 which correspond to the nucleotide positions 87 to 109 and 530 to 550 of the m5C-MTase gene of FV3, respectively. The DNA nucleotide sequence of the PCR product was determined by direct cycle sequencing. The alignment of the deduced amino acid sequence derived from the PCR product and the m5C-MTase protein of FV3 revealed a homology of 55.4% identity and 29.1% similarity. The amino acid sequence which was found to be significantly homologous to the amino acid sequence deduced from the nucleotide sequence of the PCR product was located at the amino acid position 37 to 175 of the m5C-MTase of FV3 indicating the specificity of the amplified PCR product. The DNA nucleotide sequence of the LCDV-1 genome corresponding to the 5' and 3' termini of the m5C-MTase gene was determined by primer walking. The locus of the m5C-MTase gene of LCDV-1 was identified within the EcoRI DNA fragment G of LCDV-1 (7.9 kbp; 0.947 to 0.034 map units). The m5C-MTase gene of LCDV-1 comprises 684 nucleotides coding for a putative protein of 228 amino acid residues. A high degree of amino acid sequence homology (53.3% identity and 25.8% similarity) was detected between the m5C-MTase of LCDV-1 and FV3.

This report describes a new technique for obtaining cerebrospinal fluid from the living mouse (SJL/NBom) in a model of herpes simplex virus encephalitis which is also applicable to other mouse models. The puncture technique was performed in living animals which had been infected with Herpes Simplex Virus Type I strain F in the living animal. The cisterna magna was micro-surgically prepared: The neck muscles were dissected microscopically down to the dura which subsequently was punctured by a glass micropipette. This newly developed minimally invasive technique was performed in a group of living animals (n = 20) and results compared with those of a second group of perfusion fixed animals (n = 20). For the first time, repeated cerebrospinal fluid punctures of individual, living animals are possible. This is of great value for the assessment of new therapeutic and diagnostic strategies in experimental research using mouse models. In addition, this refined methodology significantly reduces the number of experimental animals.

The complete DNA nucleotide sequence of the EcoRI DNA fragment N (0.235 to 0.258 viral map units) of equine herpes virus type 2 (EHV-2) strain T400/3 was determined. This DNA fragment comprises 4237 bp with a base composition of 55.23% G+C and 44.77% A+T. Nineteen open reading frames (ORFs) of 50-287 amino acid (aa) residues were detected. ORF number 10 is located between the nucleotide position 2220 and 2756 coding for a protein of 179 amino acid residues. This protein shows significant homology to the cytokine synthesis inhibitory factor (CSIF; interleukin 10) of human (76.4%) and mouse (68.5%), and to the Epstein-Barr virus (EBV) protein BCRF1 (70.6%). The existence of an interleukin 10 (IL-10) analogous gene within the genome of the EHV-2 was confirmed by screening the genome of nine EHV-2 strains using specific oligonucleotide primers corresponding to the 5' and 3' region of this particular gene by polymerase chain reaction. In all experiments an 870 bp DNA product was amplified. The specifity of the amplified DNA fragments obtained from individual EHV-2 strains was confirmed by DNA-DNA hybridization experiments. The DNA sequence analysis of the amplified DNA products of the EHV-2 strain LK was carried out. This analysis revealed the identity of the corresponding IL-10 gene (540 bp) of this strain to the IL-10 gene of EHV-2 strain T400/3. The presented data indicate that the EHV-2 genome harbors a viral interleukin 10-like gene. This is further evidence that the IL-10 gene can be present in the genomes of members of the Herpesviridae family.

To analyze the vif antibody response in individuals infected with the human immunodeficiency virus type 1 (HIV-1) and to determine antigenic epitopes on the vif protein, 104 HIV-1+ sera were screened for reactivity with a recombinant vif protein; 30 (28.8%) of these sera recognized the recombinant vif protein in immunoblot and were employed, together with 17 HIV-1/vif-negative control sera, in an enzyme immunoassay (EIA)-based epitope scanning assay with 183 overlapping decapeptides that covered the complete amino acid sequence of the HIV-1 vif protein (strain BH10). Of the 30 HIV-1/vif+ sera, 87% reacted with decapeptides comprising the two following epitopes: IEWRKKRY (vif amino acids 87-94) or DRWNKPQ (vif amino acids 172-178). The two epitopes were 89% and 100% conserved among different HIV-1 strains and their antigenicity could be confirmed by computer-assisted predictions of vif antigenic determinants. All the sera reactive with recombinant vif protein and with vif peptides originated from patients in CDC stages III or IV. Two murine anti-vif monoclonal antibodies reacted only with the seven C-terminal amino acids of the vif protein (SHTMNGH), which were not recognized by any of the human sera. Our results may be useful for further studies of vif seroreactivity and for the production of anti-vif mono- or polyclonal antibodies using vif peptides.