Response to and efficacy of vaccination against eastern equine encephalomyelitis virus in emus.

OBJECTIVE: To evaluate humoral immune responses of emus vaccinated with commercially available equine polyvalent or experimental monovalent eastern equine encephalomyelitis (EEE) virus and western equine encephalomyelitis (WEE) virus vaccines and to determine whether vaccinated emus were protected against challenge with EEE virus. DESIGN: Cohort study. ANIMALS: 25 emus. PROCEDURE: Birds were randomly assigned to groups (n = 5/group) and vaccinated with 1 of 2 commercially available polyvalent equine vaccines, a monovalent EEE virus vaccine, or a monovalent WEE virus vaccine or were not vaccinated. Neutralizing antibody responses against EEE and WEE viruses were examined at regular intervals for up to 9 months. All emus vaccinated with the equine vaccines and 2 unvaccinated control birds were challenged with EEE virus. An additional unvaccinated bird was housed with the control birds to assess the possibility of contact transmission. RESULTS: All 4 vaccines induced detectable neutralizing antibody titers, and all birds vaccinated with the equine vaccines were fully protected against an otherwise lethal dose of EEE virus. Unvaccinated challenged birds developed viremia (> 10(9) plaque-forming units/ml of blood) and shed virus in feces, oral secretions, and regurgitated material. The unvaccinated pen-mate became infected in the absence of mosquito vectors, presumably as a result of direct virus transmission between birds. CONCLUSIONS AND CLINICAL RELEVANCE: Results indicate that emus infected with EEE virus develop a high-titer viremia and suggest that they may serve as important virus reservoirs. Infected emus shed EEE virus in secretions and excretions, making them a direct hazard to pen-mates and attending humans. Commercially available polyvalent equine vaccines protect emus against EEE virus infection.

Risk factors for West Nile virus infection and meningoencephalitis, Romania, 1996.

In 1996, an epidemic of 393 cases of laboratory-confirmed West Nile meningoencephalitis occurred in southeast Romania, with widespread subclinical human infection. Two case-control studies were performed to identify risk factors for acquiring infection and for developing clinical meningoencephalitis after infection. Mosquitoes in the home were associated with infection (reported by 37 [97%] of 38 asymptomatically seropositive persons compared with 36 [72%] of 50 seronegative controls, P<.01) and, among apartment dwellers, flooded basements were a risk factor (reported by 15 [63%] of 24 seropositive persons vs. 11 [30%] of 37 seronegative controls, P=.01). Meningoencephalitis was not associated with hypertension or other underlying medical conditions but was associated with spending more time outdoors (meningoencephalitis patients and asymptomatically seropositive persons spent 8.0 and 3.5 h [medians] outdoors daily, respectively, P<.01). Disease prevention efforts should focus on eliminating peridomestic mosquito breeding sites and reducing peridomestic mosquito exposure.

Clinical, virological and serological responses of donkeys to intranasal inoculation with the KY-84 strain of equine arteritis virus.

The clinical, virological and serological responses of seven female donkeys (Equus asinus) to inoculation with the KY-84 strain of equine arteritis virus (EAV), a strain that causes moderate to severe clinical signs in horses, was investigated. In the donkeys, the only clinical signs observed were fever (mainly 3-9 days after inoculation), mild depression in four animals, and a slight nasal or ocular discharge in three. All of the donkeys became infected with EAV as shown by recovery of the virus for periods of up to 14 days from the nasopharynx and buffy coat and, in three out of four donkeys sampled, from the cervix or vagina. Virus replication in the donkey appeared to mirror that previously described for the horse. The donkeys had "sero-converted" to EAV by the 10th day after inoculation. Additional studies are needed to obtain a better understanding of the pathogenesis of EAV in donkeys.

Herpes simplex virus type 1 DNA cleavage and encapsidation require the product of the UL28 gene: isolation and characterization of two UL28 deletion mutants.

The herpes simplex virus type 1 UL28 gene contains a 785-amino-acid open reading frame that codes for an essential protein. Studies with temperature-sensitive mutants which map to the UL28 gene indicate that the UL28 gene product (ICP18.5) is required for packaging of viral DNA and for expression of viral glycoproteins on the surface of infected cells (C. Addison, F. J. Rixon, and V. G. Preston, J. Gen. Virol. 71:2377-2384, 1990; B. A. Pancake, D. P. Aschman, and P. A. Schaffer, J. Virol. 47:568-585, 1983). In this study, we describe the isolation of two UL28 deletion mutants that were constructed and propagated in Vero cells transformed with the UL28 gene. The mutants, gCB and gC delta 7B, contained deletions of 1,881 and 537 bp, respectively, in the UL28 gene. Although the mutants synthesize viral DNA, they fail to form plaques or produce infectious virus in cells that do not express the UL28 gene. Transmission electron microscopy and Southern blot analysis demonstrated that both mutants are defective in cleavage and encapsidation of viral DNA. Analysis by cell surface immunofluorescence showed that the UL28 gene is not required for expression of viral glycoproteins on the surface of infected cells. A rabbit polyclonal antiserum was made against an Escherichia coli-expressed Cro-UL28 fusion protein. This antibody reacted with an infected-cell protein having an apparent molecular mass of 87 kDa. The 87-kDa protein was first detected at 6 h postinfection and was expressed as late as 24 h postinfection. No detectable UL28 protein was synthesized in gCB- or gC delta 7B-infected Vero cells.

Basal forebrain and anterior thalamic contributions to acetylcholinesterase activity in granular retrosplenial cortex of rats.

Histochemical studies demonstrate that granular retrosplenial cortex (cortical areas 29b and c) of the adult rat displays a characteristic laminar pattern of acetylcholinesterase (AChE) activity. While some AChE-positive axons are found in all cortical layers, most intense staining occurs in two bands that correspond to layers I and III. The present studies were directed toward identifying the neural systems underlying this AChE activity. Unilateral electrolytic or excitatory amino acid induced lesions of the basal forebrain, including the nucleus of the diagonal band, result in reductions of AChE staining throughout ipsilateral granular retrosplenial cortex; particularly noteworthy are the reductions in layer I and the deeper cortical layers. AChE staining remains in superficial layer I and in layer III. Placement of lesions in the anterior thalamus, including all of the anterior dorsal nucleus, results in reduction of AChE histochemical staining in the outer part of layer I and especially in layer III. Staining remains in much of layer I and in the deepest band of layer III. Placement of electrolytic lesions in the hypothalamus or the midbrain tegmentum produce no detectable change in the pattern of AChE in retrosplenial cortex. These results indicate that AChE activity in granular retrosplenial cortex is found primarily within afferent axons from the basal forebrain system and from anterior dorsal thalamus, and these two systems of afferents display distinct laminar patterns of termination.

Nucleotide sequence and transcriptional studies of the vaccinia virus KpnI I DNA fragment.

The nucleotide sequence of the vaccinia virus (VV) KpnI I DNA fragment has been determined. This central, highly conserved portion of the VV genome corresponds to the right portion of the HindIII E, all of the HindIII O and P, and the left portion of the HindIII I DNA fragments. Computer-assisted analysis of this data indicated the presence of five tandemly oriented, leftward-reading open reading frames (ORFs) I-4, I-3, I-2, I-1, and O-1, with the I-4 ORF being an immediate early gene encoding the large M1 subunit of VV ribonucleotide reductase. Transcriptional analyses suggested that the I-3 and O-1 genes were constitutive genes, being expressed both before and after viral DNA synthesis. The I-1 and I-2 genes were late genes, expressed only after the initiation of viral DNA synthesis. Cell-free translation was used to confirm that the I-3, I-1, and O-1 ORFs were bonafide messages encoding proteins with molecular weights of 30, 35, and 71 kD, respectively. When the predicted amino acid sequences of the proteins encoded by the I-3, I-2, I-1, and O-1 genes were compared to the Genbank data base, no significant alignments were detected. Therefore, the biological functions of these proteins in the VV life cycle remain to be established.

Nucleotide sequence and molecular genetic analysis of the large subunit of ribonucleotide reductase encoded by vaccinia virus.

We have mapped the vaccinia virus (VV) gene encoding the large subunit of ribonucleotide reductase (VV M1) within the HindIII I restriction fragment by using an oligonucleotide probe. Nucleotide sequencing revealed a 2340-bp open reading frame (orf), 1-3, whose amino acid sequence is highly homologous to the mouse M1 protein. The 1-3 gene was expressed as an immediate-early gene product, being transcribed in a leftward direction into a 2.7-kb polyadenylated transcript. Hybrid-selected translation of cycloheximide-amplified immediate-early viral RNA demonstrated that this mRNA encoded an 86-kd protein, which agrees with the expected size of the reductase large subunit. The 5'- and 3'-boundaries of the 1-3 transcriptional unit were determined by primer extension and S1-nuclease analysis, respectively, and shown to contain sequence elements typical of other VV early genes. Surprisingly, the predicted amino acid sequence of the VV enzyme subunit shares 72.5% homology with the mouse large subunit, M1.