Ovis Strains Research Articles

Molecular epidemiology of Corynebacterium pseudotuberculosis isolated from horses in California

Corynebacterium pseudotuberculosis biovar Equi is an important pathogen of horses. It is increasing in frequency in the United States, and is responsible for various clinical forms of infection, including external abscesses, internal abscesses of the abdominal or thoracic cavities, and ulcerative lymphangitis. The host/pathogen factors dictating the form or severity of infection are currently unknown. Our recent investigations have shown that genotyping C. pseudotuberculosis isolates using enterobacterial repetitive intergenic consensus (ERIC)-PCR is useful for understanding the evolutionary genetics of the species as well for molecular epidemiology studies. The aims of the present study were to assess (i) the genetic diversity of C. pseudotuberculosis strains isolated from horses in California, United States and (ii) the epidemiologic relationships among isolates. One hundred and seven C. pseudotuberculosis biovar Equi isolates from ninety-five horses, and two C. pseudotuberculosis biovar Ovis strains, C. pseudotuberculosis ATCC 19410T type strain and C. pseudotuberculosis 1002 vaccine strain, were fingerprinted using the ERIC 1+2-PCR. C. pseudotuberculosis isolated from horses showed a high genetic diversity, clustering in twenty-seven genotypes with a diversity index of 0.91. Minimal spanning tree showed four major clonal complexes with a pattern of temporal clustering. Strains isolated from the same horse showed identical ERIC 1+2-PCR genotype, with the exception of two strains isolated from the same animal that showed distinct genotypes, suggesting a co-infection. We found no strong genetic signals related to clinical form (including internal versus external infections). However, temporal clustering of genotypes was observed.

Corynebacterium pseudotuberculosis may be under anagenesis and biovar Equi forms biovar Ovis: a phylogenic inference from sequence and structural analysis.

BackgroundCorynebacterium pseudotuberculosis can be classified into two biovars or biovars based on their nitrate-reducing ability. Strains isolated from sheep and goats show negative nitrate reduction and are termed biovar Ovis, while strains from horse and cattle exhibit positive nitrate reduction and are called biovar Equi. However, molecular evidence has not been established so far to understand this difference, specifically if these C. pseudotuberculosis strains are under an evolutionary process.ResultsThe ERIC 1 + 2 Minimum-spanning tree from 367 strains of C. pseudotuberculosis showed that the great majority of biovar Ovis strains clustered together, but separately from biovar Equi strains that also clustered amongst themselves. Using evolutionarily conserved genes (rpoB, gapA, fusA, and rsmE) and their corresponding amino acid sequences, we analyzed the phylogenetic relationship among eighteen strains of C. pseudotuberculosis belonging to both biovars Ovis and Equi. Additionally, conserved point mutation based on structural variation analysis was also carried out to elucidate the genotype-phenotype correlations and speciation. We observed that the biovars are different at the molecular phylogenetic level and a probable anagenesis is occurring slowly within the species C. pseudotuberculosis.ConclusionsTaken together the results suggest that biovar Equi is forming the biovar Ovis. However, additional analyses using other genes and other bacterial strains are required to further support our anagenesis hypothesis in C. pseudotuberculosis.Electronic supplementary materialThe online version of this article (doi:10.1186/s12866-016-0717-4) contains supplementary material, which is available to authorized users.

Identification of Brucella ovis exclusive genes in field isolates from Argentina

Brucellosis caused by Brucella ovis is one of the most important infectious diseases of sheep. The aim of this study was to determine the presence of genes both inside and outside the specific B. ovis pathogenicity island 1 (BOPI-1) in a large collection of field isolates of B. ovis and other Brucella spp. from Argentina. The BOV_A0500 gene from B. ovis BOPI-1 was identified in all 104 B. ovis isolates studied. The BOPI-1 complete sequence was found to be conserved in 10 B. ovis strains from the collection, for which whole genome sequencing was performed. The BOV_0198 gene, which is outside BOPI-1 and considered exclusive to B. ovis, showed 90–100% identity with genomic regions of B. ovis, B. melitensis, B. abortus, B. canis, B. suis, B. microti, B. ceti and B. pinnipedialis. The results demonstrate that BOPI-1 is the only exclusive genetic region of B. ovis and marine Brucella spp. and that it is highly conserved in B. ovis field isolates from Argentina.

Microarray Analysis of Gene Expression in Rams Experimentally-Infected with the Virulent Strain of Brucella ovis

Microarray analysis of gene expression profiles of total mRNA was studied in rams experimentally-infected with the virulent strain of Brucella ovis (B. ovis). The tissues studied included reproductive organs (epididymus, testicles, ampolae, vesicular glands, bulbourethral glands) and a pool of lymph nodes (inguinal and scrotal). The microarray analysis of each tissue was done at three time points: acute infection (60 days post infection [dpi]), chronic infection phase I (120 dpi), and chronic infection phase II (240 dpi). The gene expression profiles associated to B. ovis infection were determined using the Affymetrix Bovine Genome Array and expression levels of infected and uninfected rams (control group, 0 dpi) were compared. Of the 23,000 genes analyzed in the microarrays, 139 during acute phase of infection, 930 during the chronic phase I and 744 genes from the chronic phase II were identified as Differentially Expressed Genes (DEGs). Thirty known genes and fourteen unknown genes were expressed during the three phases of infection in all tissues. The biological functions of these genes included immune cell trafficking, immunological disease, infectious disease, inflammatory disease, inflammatory response and cellular movement, and significant differences were observed at the three phases of infection and in all infected tissues. For the first analysis, 8 genes in common during the three time points and to all infected tissues were chosen for the microarray validation. Acute phase of infection supported the relevance of the ataxia telangiectasia-mutated ATMIN (ATM interactor) gene. RCHY1 (ring finger and CHY zinc finger domain containing 1), ANKFY1 (ankyrin repeat and FYVE domain containing 1) and EFEMP1 (EGF-containing fibulin-like extracellular matrix protein 1) were also highlighted by the analysis during chronic phase I of infection. ZRAB2 (Zinc finger, RAN-binding domain containing 2) was validated at chronic phase II. This study represents the first analysis of microarray gene expression in different tissues of rams infected with of B. ovis at various times during infection and many of the genes identified require further study. These results expand the knowledge of the pathogenesis of this B. ovis strain infection in rams and suggest new genes and pathways for further investigation.

Genetic diversity of Brucella ovis isolates from Rio Grande do Sul, Brazil, by MLVA16.

BackgroundOvine epididymitis is predominantly associated with Brucella ovis infection. Molecular characterization of Brucella spp. achieved by multi-locus variable number of tandem repeats (VNTR) analyses (MLVA) have proved to be a powerful tool for epidemiological trace-back studies. Thus, the aim of this study was to evaluate the genetic diversity of Brucella ovis isolates from Rio Grande do Sul State, Brazil, by MLVA16.FindingsMLVA16 genotyping identified thirteen distinct genotypes and a Hunter-Gaston diversity index of 0.989 among the fourteen B. ovis genotyped strains. All B. ovis MLVA16 genotypes observed in the present study represented non-previously described profiles. Analyses of the eight conserved loci included in panel 1 (MLVA8) showed three different genotypes, two new and one already described for B. ovis isolates. Among ten B. ovis isolates from same herd only two strains had identical pattern, whereas the four isolates with no epidemiologic information exhibited a single MLVA16 pattern each. Analysis of minimal spanning tree, constructed using the fourteen B. ovis strains typed in this study together with all nineteen B. ovis MLVA16 genotypes available in the MLVAbank 2014, revealed the existence of two clearly distinct major clonal complexes.ConclusionsIn conclusion, the results of the present study showed a high genetic diversity among B. ovis field isolates from Rio Grande do Sul State, Brazil, by MLVA16.

Evaluation of ERIC-PCR as genotyping method for Corynebacterium pseudotuberculosis isolates.

The aim of this study was to evaluate the Enterobacterial Repetitive Intergenic Consensus (ERIC-PCR) as a tool for molecular typing of C. pseudotuberculosis isolates from eight different hosts in twelve countries. Ninety-nine C. pseudotuberculosis field strains, one type strain (ATCC 19410T) and one vaccine strain (1002) were fingerprinted using the ERIC-1R and ERIC-2 primers, and the ERIC-1R+ERIC-2 primer pair. Twenty-nine different genotypes were generated by ERIC 1-PCR, 28 by ERIC 2-PCR and 35 by ERIC 1+2-PCR. The discriminatory index calculated for ERIC 1, ERIC 2, and ERIC 1+2-PCR was 0.89, 0.86, and 0.92, respectively. Epidemiological concordance was established for all ERIC-PCR assays. ERIC 1+2-PCR was defined as the best method based on suitability of the amplification patterns and discriminatory index. Minimal spanning tree for ERIC 1+2-PCR revealed three major clonal complexes and clustering around nitrate-positive (biovar Equi) and nitrate-negative (biovar Ovis) strains. Therefore, ERIC 1+2-PCR proved to be the best technique evaluated in this study for genotyping C. pseudotuberculosis strains, due to its usefulness for molecular epidemiology investigations.

Complete Genome Sequence of Mycoplasma ovis Strain Michigan, a Hemoplasma of Sheep with Two Distinct 16S rRNA Genes

We report the complete genome sequence of Mycoplasma ovis strain Michigan. Its single circular chromosome has 702,511 bp and contains 2 copies of the 16S rRNA gene, one corresponding to M. ovis and the other to “Candidatus Mycoplasma haemovis.” All housekeeping genes and the 5S-23S rRNA genes are present in single copies.

The effect of 12 successive blood passages on the virulence of Babesia ovis in splenectomized lambs: A preliminary study

Babesia ovis, the primary causative agent of ovine babesiosis, is a tick-borne hemoprotozoan parasite and causes serious losses in small ruminant livestock in tropical and subtropical regions. The current control of ovine babesiosis has been base on chemotherapy and limited tick control measures. Attenuated parasites which are valuable sources of live vaccines have been used to prevent babesiosis in cattle. There is a necessity to the studies about prevention of the disease by means of immunoprophylaxis in sheep. For this purpose, this study was planned to obtain the attenuated derivatives of a virulent B. ovis strain by successive blood passages in splenectomized lambs. A total of seventeen, six-month-old lambs, 13 of which splenectomized, 4 of which spleen intact lambs, known to be negative for the presence of hemoparasites and anti-B. ovis antibodies were used. The splenectomized lambs were used for passaging the virulent B. ovis field strain. Four intact lambs divided into two groups were used to evaluate the pathogenicity of the virulent strain and twelfth passage strain of B. ovis. Blood collected from a clinically evident case of babesiosis in the field was inoculated intravenously into the first splenectomized lamb, and then the acute clinical disease was occurred. The same process was successively applied to the rest of 12 splenectomized lambs to have the attenuated B. ovis strain. The clinical, microscopic and hematological changes after inoculation were recorded in each passage of the lambs. All of the passage lambs were exhibited the severe symptoms of the disease from the 2nd day of inoculation. The same treatment protocol including vitamin B12 and the anti-babesial drug imidocarb dipropionate (IMDP) was applied to all splenectomized lambs on the fourth or fifth day of the inoculation. Although the symptoms of the disease were less severe than those of splenectomized lambs; two intact lambs infected with the virulent strain and two intact lambs infected with the twelfth passage blood were also showed the clinical and hematological findings of the disease on the 2nd and 3rd day after inoculation. Infection-related symptoms in all intact lambs disappeared spontaneously from the 6th day of inoculation without the application of any drug. The results of the study showed that there was no change on the virulence of B. ovis through the 12 successive blood passages. Therefore, the further passages or alternative attenuation methods may be necessary to attenuate the virulent strain of B. ovis.

Brucella ovis lacking a species-specific putative ATP-binding cassette transporter is attenuated but immunogenic in rams

Ovine brucellosis caused by Brucella ovis is considered one of the most important reproductive diseases of rams worldwide. This study aimed to characterize the kinetics of infection of a ΔabcAB B. ovis mutant strain in rams. Twelve 1-year-old crossbred rams were used. Six rams were challenged with 2mL of a suspension containing 1.2×109CFU/mL of B. ovis strain ATCC25840 (wild type) by intraprepucial inoculation and additional 50μL in each conjunctival sac of a suspension containing 1.2×1010CFU/mL of the same strain. The other six rams were challenged with an equivalent number of CFU of the mutant strain ΔabcAB B. ovis through the same routes. Serum samples for serology and semen and urine samples for bacteriologic culture and PCR were collected weekly during 24 weeks. At 24 weeks post infection, tissue samples were collected for bacteriologic culture and PCR. All rams inoculated with wild type or the ΔabcAB strain seroconverted at the fourth week post infection, remaining positive up to the 16th week post infection. PCR and bacteriology demonstrated that only rams inoculated with the wild type strain shed the organism in semen and urine. Lymphocytes from rams inoculated with wild type or ΔabcAB B. ovis had significantly higher proliferation in response to B. ovis antigens when compared with unstimulated controls. Tissue bacteriology and PCR detected B. ovis in all rams challenged with the wild type strain, whereas only one ΔabcAB-infected ram had a positive iliac lymph node sample by PCR.

Differential Exoproteome Analysis of Two Corynebacterium pseudotuberculosis Biovar Ovis Strains Isolated from Goat (1002) and Sheep (C231)

Corynebacterium pseudotuberculosis is the etiologic agent of caseous lymphadenitis a chronic infectious disease affecting small ruminants. The 2D-DIGE technique was used to compare the exoproteomes of two C. pseudotuberculosis biovar ovis strains isolated from goat (strain 1002) and sheep (strain C231). Seventeen proteins differentially produced were identified here. Nine proteins appeared over-produced in the exoproteome of 1002 goat strain and 8 in that of C231 sheep strain. These proteins were related to various biological functions, such as the cell envelope, respiratory metabolism and proteolysis. This proteomic analysis revealed strain-specific exoproteins although each of the corresponding genes was found in both strain genomes. Such differential expression pattern may reflect inter-strain differences in adaptation to a specific host, in pathogenicity and or in antigenicity of this pathogenic bacterium.

Natural IS711 insertion causing omp31 gene inactivation in Brucella ovis

The present report describes an atypical Brucella ovis strain (Bo10) isolated from the epididymis and testis of an infected ram. Macroscopic and microscopic lesions characteristic for the infection, including positive Brucella immunostaining, were observed within lesions in the genital organs. Compared to other isolates, strain Bo10 required an additional day (a total of 96 hr) of incubation to form visible colonies, showed a distinct carbon source utilization profile, agglutinated only weakly with rough (R) serum, but showed a high capacity for autoagglutination. Isolate Bo10 failed to produce the 1,071-bp fragment in the outer membrane protein (omp) 31 gene-based part of the "Bruce-ladder" multiplex polymerase chain reaction system but did produce a 1,915-bp amplicon, thus presenting a profile similar to Brucella abortus. Sequence analysis of the 1,915-bp fragment revealed an 842-bp long insertion sequence (IS)711 transposon element inserted into the promoter region of the omp31 gene, immediately upstream from the ribosome binding site (-10 box/Pribnow box). Sodium dodecyl sulfate-polyacrylamide gel electrophoresis of a whole-cell lysate showed the absence in Bo10 of the approximately 31-kDa protein fragment associated with omp31. The results demonstrate a natural inactivation of omp31 and, consequently, the absence of the Omp31 protein in this B. ovis isolate. The novel location of IS711 within the genome of a naturally occurring B. ovis strain supports the hypothesis that IS711 could be an active transposon in this Brucella species.

Complete genome sequence of Corynebacterium pseudotuberculosis biovar ovis strain P54B96 isolated from antelope in South Africa obtained by rapid next generation sequencing technology.

The Actinobacteria, Corynebacterium pseudotuberculosis strain P54B96, a nonmotile, non-sporulating and a mesophile bacterium, was isolated from liver, lung and mediastinal lymph node lesions in an antelope from South Africa. This strain is interesting in the sense that it has been found together with non-tuberculous mycobacteria (NTMs) which could nevertheless play a role in the lesion formation. In this work, we describe a set of features of C. pseudotuberculosis P54B96, together with the details of the complete genome sequence and annotation. The genome comprises of 2.34 Mbp long, single circular genome with 2,084 protein-coding genes, 12 rRNA, 49 tRNA and 62 pseudogenes and a G+C content of 52.19%. The analysis of the genome sequence provides means to better understanding the molecular and genetic basis of virulence of this bacterium, enabling a detailed investigation of its pathogenesis.

The virB-encoded type IV secretion system is critical for establishment of infection and persistence of Brucella ovis infection in mice

Brucella spp. are gram-negative intracellular bacterial pathogens that cause chronic infections. Brucella virulence factors include a type IV secretion system (T4SS) and its lipopolysaccharide (LPS), which are essential for persistence. However, the role of the virB-encoded T4SS has not been investigated in naturally rough Brucella species such as Brucella ovis. In this study, male 6-week old BALBc mice were infected with B. ovis, Brucella abortus, and their respective ΔvirB2 mutant strains. During early infection, B. ovis and B. abortus wild type strains were similarly recovered from spleen. Interestingly, in contrast to ΔvirB2 B. abortus that was recovered at similar levels when compared to the wild type strain, the ΔvirB2 B. ovis was markedly attenuated as early as 24h post infection (hpi). The ΔvirB2 B. ovis was unable to survive and multiply in murine peritoneal macrophages and extracellularly within the peritoneal cavity at 12 and 24hpi with lower splenic colonization than the parental strain at 6, 12 and 24hpi. In contrast, wild type B. abortus and ΔvirB2 B. abortus had a similar kinetics of infection in this model. As expected, the T4SS was essential for intracellular replication of smooth and rough strains in RAW macrophages at 48hpi. These results suggest that T4SS is important for survival of B. ovis in murine model, and that a T4SS deficient B. ovis strain is cleared at earlier stages of infection when compared to a similar B. abortus mutant.

Andrological, pathologic, morphometric, and ultrasonographic findings in rams experimentally infected with Brucella ovis

Brucella ovis is considered the most important infectious cause of reproductive disorders in sheep. The disease is characterized by epididymitis, subfertility and infertility in rams. B. ovis occasionally results in abortion in ewes, as well. The aim of this study was to evaluate kinetic changes in the reproductive organs of rams experimentally infected with B. ovis. Nine rams were experimentally inoculated intrapreputially with 2mL of a suspension containing 1.2×109CFU (colony-forming units)/mL of B. ovis (strain ATCC25840). In addition, 50μL of a suspension containing 1.2×1010CFU/mL of the same B. ovis strain was inoculated into each conjunctival sac, resulting in 3.6×109CFU total per ram. Six of nine infected rams had developed clinical changes in the tail of the epididymis at 30 days post-infection (dpi), but these changes regressed in 50% of these rams. Ultrasound demonstrated an increase in the area of the tail of the epididymis (P<0.001), reduction in the area of the testes (P<0.001), and an increased length and width of the seminal vesicles (P<0.001) during the course of infection. A sperm granuloma was diagnosed on the basis of ultrasonography findings. Microscopically, there was epididymitis, testicular degeneration, and seminal vesiculitis. Inflammatory cells were detected in the semen even before the development of epididymitis. Moreover, inflammatory cells were also found in the semen of asymptomatic rams, indicating that the presence of leukocytes in the ejaculate is a valuable method for screening potential carriers of infections in the genital tract.

A potential novel Brucella species isolated from mandibular lymph nodes of red foxes in Austria

The wild red fox ( Vulpes vulpes) is a known indicator species for natural foci of brucellosis. Here, we describe phenotypic and molecular characteristics of two atypical Brucella strains isolated from two foxes hunted 2008 in Eastern Austria. Both strains agglutinated with monospecific anti- Brucella A serum and were positive in ELISA with monoclonal antibodies directed against various Brucella lipopolysaccharide epitopes. However, negative nitrate reductase- and negative oxidase-reaction were atypical traits. Affiliation to the genus Brucella was confirmed by 16S rRNA gene sequencing and by detection of the Brucella specific insertion element IS 711 and gene bcsp31 using real-time PCR. Both fox strains showed identical IS 711 Southern blot profiles but were distinct from known brucellae. The number of IS 711 copies detected was as high as found in B. ovis or marine mammal Brucella strains. Molecular analyses of the recA and omp2a/b genes suggest that both strains possibly represent a novel Brucella species.

Fatal bovine anaplasmosis in a herd with new genotypes of Anaplasma marginale, Anaplasma ovis and concurrent haemoplasmosis

Haematological and molecular analysis of blood samples was carried out during an outbreak of bovine anaplasmosis in Hungary. Acute disease was observed in five animals, two of which died. Anaplasma-carrier state was diagnosed in 69 (92%) of cattle. Further evaluation of 24 blood samples revealed concurrent infections with Mycoplasma wenyonii and ‘ Candidatus M. haemobos’ in 22 and 21 animals, respectively. In addition, two cows were identified with rickettsaemia. Regarding molecular investigation of potential hard tick vectors, Haemaphysalis inermis and Dermacentor marginatus males collected from the animals were PCR-negative. However, in one pool (out of 18) of Ixodes ricinus males, and in six pools (out of 18) of D. reticulatus males the msp4 gene of Anaplasma marginale was detected. In the same I. ricinus pool Anaplasma ovis was also identified. All ticks were negative for haemoplasmas. Anaplasma sequences yielded 97–99% homology to sequences deposited in the Genbank. This is the first report of fatal bovine anaplasmosis associated with divergent A. marginale genotypes and concurrent ‘ Candidatus M. haemobos’ infection, as well as of an A. ovis strain in ticks collected from cattle.

Human Anaplasmosis andAnaplasma ovisVariant

Human Anaplasmosis and<i>Anaplasma ovis</i>Variant

Characterization of Anaplasma phagocytophilum and A. ovis infection in a naturally infected sheep flock with poor health condition

Anaplasma species are transmitted by ticks and cause diseases in humans and animals. These pathogens infect sheep, an economically important domestic animal worldwide. The current study was designed to characterize in 200 animals the infection with Anaplasma phagocytophilum and Anaplasma ovis and the genetic diversity of A. ovis strains collected from a naturally infected sheep flock with poor health condition. Sheep had 98% seroprevalence to Anaplasma spp. antibodies. PCR results confirmed the presence of A. phagocytophilum and A. ovis DNA in 11.5% and 37% of the sheep, respectively. Concurrent infections were detected in 6.5% of the sheep. Seventy-one adult ticks were collected from 45 sheep with infestations ranging from one to 15 ticks per animal. The analysis of A. ovis msp4 sequences demonstrated a previously unreported polymorphism for this pathogen with 17 different haplotypes in infected sheep. These results demonstrated that, although A. ovis msp4 haplotypes may be less variable when compared with Anaplasma marginale and A. phagocytophilum strains on a global scale, genetic polymorphisms occur in this locus in strains obtained from an infected sheep flock with poor health condition.

The feasibility of using antigens prepared with rough Brucella strains for diagnosis of canine brucellosis.

Clinical diagnosis of canine brucellosis is not sensitive enough and a negative blood culture cannot rule out the disease. Indirect methods of serological testing such as agar gel immunodiffusion (AGID), rapid slide agglutination test (RSAT) and indirect enzyme linked immunoassay (IELISA) are preferred for routine diagnosis. Since Brucella canis shares antigenic components with the Brucella ovis and Brucella abortus RB51 strain, it would seem that either strain could be used as antigen. We present data on AGID and IELISA tests using the B. ovis antigen, RSAT and IELISA using the B. canis antigen and IELISA using the B. abortus RB51 antigen. The cut-off values were adjusted by the ROC analysis; the IELISA-B. ovis cut-off value was 23 (%P) and the IELISA-B. abortus RB51, 24 (%P), with 100% sensitivity and 98.8% specificity. RSAT detected 100% of positive cases, while AGID was less sensitive. The sera from dogs treated with antibiotic showed that %P correlated well with the clinical course. Sera from dogs presumptively infected with B. suis were negative in all tests performed with the rough Brucella strains. RSAT is a very sensitive screening test and IELISA-B. canis, B. ovis and B. abortus RB51 could be used as confirmatory tests, since they show good specificity and sensitivity.

Interferon-γ Prevents Death of Bystander Neurons during CD8 T Cell Responses in the Brain

T cells restricted to neurotropic viruses are potentially harmful as their activity may result in the destruction of neurons. In the Borna disease virus (BDV) model, antiviral CD8 T cells entering the brain of infected mice cause neurological disease but no substantial loss of neurons unless the animals lack interferon-gamma (IFN-gamma). We show here that glutamate receptor antagonists failed to prevent BDV-induced neuronal loss in IFN-gamma-deficient mice, suggesting that excitotoxicity resulting from glutamate receptor overstimulation is an unlikely explanation for the neuronal damage. Experiments with IFN-gamma-deficient mice lacking eosinophils indicated that these cells, which specifically accumulate in the infected brains of IFN-gamma-deficient mice, are not responsible for CA1 neuronal death. Interestingly, BDV-induced damage of CA1 neurons was reduced significantly in IFN-gamma-deficient mice lacking perforin, suggesting a key role for CD8 T cells in this pathological process. Specific death of hippocampal CA1 neurons could be triggered by adoptive transfer of BDV-specific CD8 T cells from IFN-gamma-deficient mice into uninfected mice that express transgene-encoded BDV antigen at high level in astrocytes. These results indicate that attack by CD8 T cells that cause the death of CA1 neurons might be directed toward regional astrocytes and that IFN-gamma protects vulnerable CA1 neurons from collateral damage resulting from exposure to potentially toxic substances generated as a result of CD8 T cell-mediated impairment of astrocyte function.