A case report of long-delayed diagnosis of pseudorabies virus encephalitis with endophthalmitis: lessons from metagenomic next generation sequencing.

BACKGROUND: Pseudorabies virus (PRV) was thought to only infect animals. Recent studies have shown that it can also infect human. CASE PRESENTATION: We report a case of pseudorabies virus encephalitis and endophthalmitis, diagnosed 89 days after onset, confirmed with intraocular fluid metagenomic next generation sequencing (mNGS) after the result of two cerebrospinal fluid (CSF) mNGS tests were negative. Although treatment with intravenous acyclovir, foscarnet sodium, and methylprednisolone improved the symptoms of encephalitis, significant diagnostic delay resulted in permanent visual loss. CONCLUSIONS: This case suggests that pseudorabies virus (PRV) DNA in the intraocular fluid may have a higher positivity than that in the CSF. PRV may persist in the intraocular fluid for an extended period and may thus require extended antiviral therapy. Patients with severe encephalitis and PRV should be examined with the focus on pupil reactivity and light reflex. A fundus examination should be performed in patients with a central nervous system infection, specifically, those in a comatose state, to help reduce eye disability.

Abnormal spermatogenesis and reduced fertility in transgenic mice expressing the immediate-early protein IE180 of pseudorabies virus.

Transcription factors of alphaherpesviruses not only control the expression of their own viral genes, but also influence the gene expression of mammalian cells. In the course of breeding of the transgenic mouse line (TgIE96) expressing the immediate-early protein IE180 of pseudorabies virus belonging to the subfamily Alphaherpesvirinae, we found that TgIE96 male mice suffered from severe breeding difficulties. Testes of TgIE96 were smaller than that of non-transgenic littermates and abnormal spermatogenesis such as morphological, numerical and functional anomalies of spermatozoa were found in the transgenic mouse line. Expression of IE180 was detected in the germ cells at all stages, especially spermatocytes, and fewer Sertoli cells. In addition, expression of IE180 was also detected in the germinal cells of C57BL/6 mice inoculated with PRV into their testes. These results suggest that IE180 of PRV induces male infertility by abnormal spermatogenesis, which effect morphological, numerical, and functional anomalies of spermatozoa, in transgenic mice.

Organ cross talk modulates pelvic pain.

Interstitial cystitis (IC) is a chronic bladder inflammatory disease of unknown etiology that is often regarded as a neurogenic cystitis. IC is associated with urothelial lesions, voiding dysfunction, and pain in the pelvic/perineal area, and diet can exacerbate IC symptoms. In this study, we used a murine neurogenic cystitis model to investigate the development of pelvic pain behavior. Neurogenic cystitis was induced by the injection of Bartha's strain of pseudorabies virus (PRV) into the abductor caudalis dorsalis tail base muscle of female C57BL/6J mice. Infectious PRV virions were isolated only from the spinal cord, confirming the centrally mediated nature of this neurogenic cystitis model. Pelvic pain was assessed using von Frey filament stimulation to the pelvic region, and mice infected with PRV developed progressive pelvic pain. Pelvic pain was alleviated by 2% lidocaine instillation into either the bladder or the colon but not following lidocaine instillation into the uterus. The bladders of PRV-infected mice showed markers of inflammation and increased vascular permeability compared with controls. In contrast, colon histology was normal and vascular permeability was unchanged, suggesting that development of pelvic pain was due only to bladder inflammation. Bladder-induced pelvic pain was also exacerbated by colonic administration of a subthreshold dose of capsaicin. These data indicate organ cross talk in pelvic pain and modulation of pain responses by visceral inputs distinct from the inflamed site. Furthermore, these data suggest a mechanism by which dietary modification benefits pelvic pain symptoms.

Brain lesions in pigs dually infected with porcine reproductive and respiratory syndrome virus and pseudorabies virus.

Four pigs were inoculated with an aerosol containing porcine reproductive and respiratory syndrome virus (PRRSV) followed 14 days later by inoculation with pseudorabies virus (PRV). The four dually infected pigs showed severe clinical signs, and one died on day 6 after infection with PRV. As demonstrated previously, the clinical disease was much more severe than that produced by either virus alone. All four dually infected pigs developed severe non-suppurative encephalitis, two had tonsillitis, two had necrotizing bronchiolitis, and one had lymphadenitis. The distribution of lesions corresponded closely with the detection of intranuclear inclusion bodies and PRV antigen. High numbers of TUNEL-positive cells detected in the thymus were associated with thymic atrophy.

Immunogenicity of a recombinant pseudorabies virus expressing ORF1-ORF2 fusion protein of porcine circovirus type 2.

Porcine circovirus type 2 (PCV2) is associated with post-weaning multisystemic wasting syndrome (PMWS). Pseudorabies (PR) is also an important infectious disease in swine and sometimes co-infect with PCV2. An attenuated pseudorabies virus (PRV) has been successfully used as a vector for live viral vaccines. In this study, a recombinant PRV expressing ORF1-ORF2 fusion protein of PCV2 was constructed and its immunogenicity was tested in mice and pigs. The ORF1 and partial ORF2 gene of PCV2 Yu-A strain were amplified by PCR and inserted into a transfer vector. The recombinant transfer plasmid was co-transfected with the EcoRI digested genome of vector virus (PRV TK-/gE-/LacZ+) into IBRS-2 cells. The recombinant pseudorabies virus PRV-PCV2 was purified by plaque purification and identified by PCR and Southern blotting. Expression of the ORF1-ORF2 fusion protein by the recombinant PRV-PCV2 virus was demonstrated by Western blotting analysis. The growth properties of the recombinant virus in cells were similar to that of the parent vector virus. In animal experiments, PRV-PCV2 elicited strong anti-PRV and anti-PCV2 antibodies in Balb/c mice as indicated by PRV-neutralizing assay, anti-PCV2 ELISA and PCV2 specific lymphocyte proliferation assay, respectively. And PRV-PCV2 immunization protected mice against a lethal challenge of a virulent PRV Ea strain. In pigs, PRV-PCV2 elicited significant immune response towards PRV and PCV2 as indicated by PRV-ELISA, PRV neutralizing assay and PCV2 specific lymphocyte proliferation assay, respectively. This is a first step toward the development of a potential candidate divalent vaccine against PRV and PCV2 infections.

CNS induced neurogenic cystitis is associated with bladder mast cell degranulation in the rat.

PURPOSE: To determine if bladder mast cell degranulation is involved in the genesis of neurogenic cystitis induced by pseudorabies virus (PRV) invasion of the central nervous system (CNS). MATERIALS AND METHODS: Rats received a total of 4 x 106 plaque forming units (pfu) of PRV-Bartha in the abductor caudalis dorsalis (ACD) muscle. Granulated bladder mast cells per mm2 of bladder tissue and urine histamine content were monitored as the cystitis developed over the next few days. In a subgroup of rats, intravesical resiniferatoxin was used to remove capsaicin-sensitive sensory bladder afferents, while another subgroup was pretreated with a mast cell degranulator. RESULTS: PRV injection into the ACD muscle leads to neurogenic cystitis. Histamine levels were elevated in the urine of virus injected rats before any behavioral or microscopical signs of cystitis were present. When the cystitis became clinically manifest, urine histamine returned to control levels, and the number of granulated mast cells dropped significantly. Rats in which capsaicin-sensitive afferents had been removed did not show any signs of cystitis, or increase in urine histamine, or change in the number of granulated mast cells. Pretreatment of animals with a mast cell degranulator completely prevented the appearance of cystitis without altering the CNS disease. CONCLUSION: These results provide further evidence that mast cells are involved in neurogenic cystitis induced by changes in CNS activity.

Simultaneous serological evidence of Actinobacillus pleuropneumoniae, PRRS, Aujeszky's disease and influenza viruses in Spanish finishing pigs.

A total of 198 pigs with tachypnoea and temperature >/= 40 degrees C were selected on a Spanish finishing unit, and their sera were examined for antibodies to Actinobacillus pleuropneumoniae (App), porcine reproductive and respiratory syndrome virus (PRRSV), Aujeszky' disease virus (ADV), and swine influenza virus (SIV). Eighty-nine point nine per cent of the pigs were seropositive to App, 88.6 per cent to PRRS, 73.0 per cent to ADV, and 30.6 per cent to SIV. Thirty-one pigs (15.6 per cent) were seropositive for App, PRRSV, ADV and SIV, and only one (0.5 per cent) was seronegative for all. Statistical association was assessed for dual infections but it was not found in any case (P > 0.05). Other parameters (dyspnoea, nasal discharge and coughing) were also recorded, and no significant associations between them and the presence of antibodies against any of the four infections was found.

Effects of mild stress on the immune response against pseudorabies virus in mice.

Stress is a recognised problem in intensive pig husbandry, which might lead to changes in immune reactivity. To study the effect of stress on the development of an anti-viral immune response, we used a murine model in which mice were immunized with an attenuated strain of pseudorabies virus (PRV). The effect of two stress treatments, both relevant to intensive pig husbandry, on the development of the specific immune response against PRV was investigated. The stress treatments consisted of restraint, social isolation, and transport and they differed in predictability. The specific immune response against PRV, which developed in the draining lymph nodes, was measured by a lymphocyte proliferation assay and cytokine production assays. Our results showed that the unpredictable stress treatment had no effect on the development of the immune response against PRV in mice, whereas the predictable stress treatment actually hastened the immune response.

Experimental dual infection of cesarean-derived, colostrum-deprived pigs with Mycoplasma hyopneumoniae and pseudorabies virus.

To determine whether pseudorabies virus (PRV) infection increases the severity of pneumonia by Mycoplasma hyopneumoniae, 18, 10-week-old Cesarean-derived, colostrum-deprived pigs were randomly assigned to 3 groups of 6 pigs each. Pigs in groups A and C were inoculated intranasally with M. hyopneumoniae at 10-week-old. At 11-week-old, pigs in groups B and C were inoculated intranasally with PRV. All pigs were initially seronegative for M. hyopneumoniae and PRV. Three pigs of each group were euthanized at 12-week-old, and remaining pigs at 14-week-old. At necropsy, gross lesions in the lung were observed in the pigs of groups A and C. On post-inoculation-week (PIW) 2 with M. hyopneumoniae (at 12-week-old), lung lesions were recognized in one of the 3 pigs in group A and all the pigs in group C. The mean percentage of the lung lesions were 0.1% in group A and 9.8% in group C. M. hyopneumoniae was isolated from broncho-alveolar lavage fluids (BALF) of pigs in group A with titer of 10(2) to 10(3) CCU/0.2 ml and in group C with titer of 10(5) to 10(6) CCU/0.2 ml. On PIW 4 (at 14-week-old), lung lesions were observed in all the pigs in groups A and C, and the mean percentage of the lung lesions were 8.3% in group A and 17.2% in group C. M. hyopneumoniae was isolated from BALF in group A with titer of 10(4) to 10(7) CCU/0.2 ml and in group C with titer of 10(6) to 10(7) CCU/0.2 ml. PRVs were isolated from nasal swab and tissue samples in groups B and C. After inoculation, antibody against M. hyopneumoniae was detected in groups A and C, and against PRV in groups B and C. Under the present experimental conditions, PRV infection appear to have effect on the severity of experimentally induced acute mycoplasmal pneumonia in young pigs.

Serological studies on porcine Pneumocystis carinii pneumonia: kinetics of the antibody titers in swine herds and the association of porcine reproductive and respiratory syndrome virus infection.

Serological titers to Pneumocystis carinii (Pc) and porcine reproductive and respiratory syndrome virus (PRRSV) were measured on a herd with epidemic Pc pneumonia (case herd) and two comparison herds, by an indirect fluorescent-antibody technique. In the case herd, the geometric mean titer (GMT) for Pc were 1:80 in pigs 1 week old, 1:10 in pigs 5 weeks old, and 1:80 to 1:190 in pigs over 6 weeks old. GMTs for PRRSV were > 1:145 in most of age groups over 7 weeks old. In comparison herds, Pc and PRRSV antibody titers were low in weanling pigs. The results clarified the kinetics of antibodies to Pc and concurrent infection of PRRSV in the case herd.

Bilateral encephalomalacia in a pig related to latent Aujeszky's disease virus infection.

A case of bilateral encephalomalacia in a pig related to latent Aujeszky's disease virus infection is reported. The pig was experimentally inoculated with the NIA-3 strain and survived the infection after showing intense central nervous system disease. Abnormal behaviour was observed up to the date of death. The pig was demonstrated to be latently infected with the virus by polymerase chain reaction (PCR). The main microscopic lesion was a bilateral encephalomalacia which involved structures related to the limbic system. A complete description of lesions observed and their relation to abnormalities shown by the pig are exposed.

Cell-free and cell-associated viremia in pigs after oronasal infection with Aujeszky's disease virus.

Nine pigs were examined for the presence of viremia during the first week after oronasal inoculation of 10(8.0) TCID50 Aujeszky's disease virus (ADV). Blood was taken at 1, 2, 3, 5 and 7 days post inoculation (PI) and the presence of cell-free ADV in plasma and of ADV-infected mononuclear cells was examined by titration and by cocultivation with permissive cells, respectively. The mononuclear cells of 6 of the 9 pigs, collected at 3 and 5 days PI were further separated into subpopulations of enriched monocytes and enriched lymphocytes. Both subpopulations were cocultivated. Nasal secretions were collected from 4 of the 9 pigs for the determination of virus titers and interferon concentrations. Both infected mononuclear cells and cell-free ADV were demonstrated in 5 pigs, infected mononuclear cells only were found in 2 pigs, and neither cell-associated or cell-free ADV were detected in 2 pigs. Two of the 7 viremic animals were positive on one single day, 3 on 2 days, 1 on 3 days and 1 on 4 days. The number of infected cells was approximately 5 times higher in monocytes than in lymphocytes. The highest virus titers were present in those nasal fluids with the lowest alpha-interferon concentration. A correlation between the titer of locally produced ADV in the nose and the presence of a viremia was not found. In conclusion, we can state that a viremia regularly occurs under both cell-free and cell-associated form after an oronasal inoculation of ADV and that monocytes are the most susceptible mononuclear cells.

Invasion and spread of single glycoprotein deleted mutants of Aujeszky's disease virus (ADV) in the trigeminal nervous pathway of pigs after intranasal inoculation.

The purpose of the study was to evaluate the role which non-essential envelope glycoproteins play in the neuroinvasion and neural spread of ADV. The invasion and spread in the trigeminal nervous pathway with the Ka strain of ADV and its single deletion mutants Ka gI-, Ka gp63- and Ka gIII- were examined after intranasal inoculation in neonatal pigs by virus isolation and immunocytochemistry. Evaluation was performed in the nasal mucosa, trigeminal ganglion (1st neuronal level), ponsmedulla (2nd neuronal level) and thalamus-cerebellum (3rd neuronal level). The Ka gIII- mutant invaded up to the 3rd neuronal level of the trigeminal pathway and spread in a similar way to the parental Ka strain. The Ka gp63- mutant invaded up to the 3rd neuronal level but the spread of this mutant was impaired at all the neuronal levels. The Ka gI- mutant was least neuroinvasive and reached only up to the 2nd neuronal level. The results showed that glycoproteins gI and gp63 play a role in the invasion and spread of ADV in the nervous system. However, the gI glycoprotein appears to be the most important for neuroinvasion and neural spread of ADV in pigs. Therefore, gI deleted vaccines may be considered to be safer with respect to the neuroinvasion than vaccines carrying single deletions of other non-essential envelope glycoproteins.

Pneumonia in pigs infected with pseudorabies virus and Haemophilus parasuis serovar 4.

Six HPCD (hysterectomy-produced, colostrum-deprived) pigs were inoculated endobronchially with pseudorabies virus (PRV) in the right caudal lobe by means of a bronchoscope. Two pigs, killed on days 5 and 7, had severe purulent pneumonia in the right caudal lobe, associated with an accidental Haemophilus parasuis serovar 4 infection. The three surviving animals were treated with antibiotics. The pigs infected with PRV had necrotizing bronchiolitis and alveolitis. PRV antigen was closely associated with necrotic foci, and was sometimes surrounded by profuse H. parasuis antigen. PRV antigen and IgG- and IgA-containing cells were also detected in bronchioalveolar lavage fluid. These results suggested that the PRV infection destroyed respiratory epithelial cells and allowed H. parasuis to proliferate in the lungs.

Mechanism of pruritus and peracute death in mice induced by pseudorabies virus (PRV) infection.

Mechanisms of postinfectious pruritus and peracute death in mice by pseudorabies virus (PRV) were investigated by inoculating the Yamagata-S81 strain of PRV peripherally or intracerebrally into 4-week-old ICR and BALB/c mice. Clinical signs developed most rapidly in mice inoculated intracerebrally, with intermediate speed in mice inoculated intraocularly, and slowly in mice inoculated subcutaneously. Since intraocularly inoculated mice showed an acute reaction and this is considered a peripheral route, the distribution of viral antigens in the nervous system of intraocularly inoculated mice was examined immunohistologically. Viral antigens were mainly detected along the trigeminal and the oculomotor nerves, but neither necrosis nor an inflammatory response was observed in these areas. The infectious virus was efficiently recovered from the viral antigen-positive tissues. In the pruritic skin lesions, viral antigens were not observed. These findings indicate that the main route of viral spread in intraocularly inoculated mice is the trigeminal and oculomotor nerves and that the virus in the trigeminal nerve may trigger pruritus.

Pneumonia in pigs inoculated with pseudorabies virus by means of a bronchoscope.

Pigs inoculated endobronchially (EB) with 2 ml of virus suspension containing 10(4) TCID50 per ml of the YS-81 strain of pseudorabies virus (PRV), by means of a bronchoscope, all developed viral pneumonia. No pneumonic lesions were observed in intranasally inoculated pigs. Macroscopical and microscopical lesions were localized to the middle to caudal parts of the right caudal lobe and were closely associated with the site at which the inoculum was deposited. PRV became attached to all types of cells and caused destruction of epithelial cells, and viral antigen persisted in the alveolar macrophages. After PRV infection, the total cell number in broncho-alveolar lavage (BAL) fluid was slightly increased and a high titre of PRV was found in the cells of BAL fluid in EB infected pigs. The findings suggest that PRV infection leads to dysfunction of alveolar macrophages before cell death is produced by virus replication.

Inoculation of pigs with Streptococcus suis type 2 alone or in combination with pseudorabies virus.

Pigs (9 [+/- 1] weeks old) were inoculated with Streptococcus suis type 2, pseudorabies virus (PRV), or both. For each pig of groups A, B, and C the inoculum of S suis was 10(9) colony-forming units. For each pig of groups A, B, and D the inoculum of PRV was 5 x 10(3) TCID50 of either PRV strain 4892 (group A, n = 9) or PRV isolate B (group B, n = 9). The PRV strain 4892 is a highly virulent strain; isolate B causes mild clinical signs of infection in inoculated pigs. Group-C pigs (n = 9) were given S suis alone, and group-D pigs (n = 3) were inoculated only with PRV isolate B. Clinical signs of infection and development of lesions were readily seen in pigs of groups A, B, and C. Duration and severity of clinical signs of disease and lesions were reduced in pigs of group C, compared with those of the other 2 groups. Lesions, such as polyarthritis and fibrinous pericarditis, were more abundant and acute in the groups of pigs given mixed challenge exposure, compared with pigs inoculated exclusively with S suis type 2 (group C). The group of pigs inoculated with PRV isolate B alone did not manifest clinical signs of disease or lesions. Average daily gain for group-C pigs was higher, compared with that of other groups; the difference was statistically significant at P less than 0.02 and P less than 0.05 for groups B and D, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Multiple neurologic deficits. Inflammatory diseases.

Certain diseases do not consistently affect a single area of the nervous system and, in fact, may cause concomitant multifocal involvement. Depending on the area of the nervous system affected, these diseases may cause a wide spectrum of neurologic problems and therefore must be included in the differential diagnosis for most problems discussed previously. Diseases causing multifocal lesions should especially be suspected when there are concomitant neurologic deficits that cannot be localized to a single site. An example would be a cat with seizures and paraparesis without thoracic limb involvement, in which case separate lesions affecting the forebrain and thoracolumbar spinal cord would be suspected. Inflammatory neurologic diseases most commonly cause lesions at multiple sites and will be discussed here. These conditions frequently cause concomitant systemic disease. Additional disease categories that may cause multifocal lesions, and are somewhat more likely to result in strict neurologic disease, are discussed in Multiple Neurologic Deficits: Non-infectious Diseases (page 440).

Prevalence of pseudorabies virus infection and associated infections in six large swine herds in Illinois.

Sera were collected from 6 large farrow-to-finish swine herds infected with pseudorabies virus (PRV) in Illinois. All herds were participating in the Large Herd Cleanup Study, a USDA-initiated project to evaluate the feasibility of eradicating pseudorabies from large farms (greater than 400 sows) by use of a combination of vaccination and management changes. Herd size ranged between 425 and 1,500 breeding females. Between April and July 1990, sera for measurement of PRV antibodies were obtained from 113 to 156 sows and 112 to 162 finishing pigs (body weight greater than 70 kg)/herd. Duplicate sera from 30 sows and 30 market-weight pigs/herd were obtained for measurement of serum antibodies to the following associated organisms: swine influenza virus, transmissible gastroenteritis virus, encephalomyocarditis virus, Actinobacillus pleuropneumoniae, Eperythrozoon suis, and 6 serovars of Leptospira interrogans. Prevalence of PRV antibodies attributable to field virus infection ranged between 53.8 and 100% for sows and between 0.7 and 97.3% for finishing pigs, as determined by the appropriate differential test for the vaccine being used on each farm. In only 1 herd, PRV seroprevalence was increased with higher sow parity. For associated infections, the risk of seropositivity attributable to PRV was not significant (for most infections) on all farms and varied among farms. Thus, pseudorabies did not appear, in general, to increase susceptibility to infection with other disease agents.